E-8 F-1 Regulation of Carbon Metabolic Genes, adhA and · PDF fileCNQ-509 JINSOO PARK 2, Hak...

305The Korean Society for Microbiology and Biotechnology

2010 International Symposium & Annual Meeting

Regulation of Carbon Metabolic Genes, adhA and ptsG, in An Adenylate Cyclase Gene Deletion Mutant of Corynebacterium Glutamicum ATCC 13032 Bindu SUBHADRA, Jong Yun HAN and Jung-Kee LEE*Department of Life Science and Genetic Engineering, Paichai University, Daejeon302-735, Korea. *Corresponding author: [email protected]

The cyclic AMP receptor protein (CRP) homologue, GlxR, controls the expression of several genes involved in the regulation of diverse physiological processes in Corynebacterium glutamicum. In silico analysishas revealed the presence of glxR binding sites upstream of genes, ptsGand adhA, encoding glucose-specific phosphotransferase system protein and alcohol dehydrogenase (ADH). However, the involvement of GlxR-cAMP complex on the expression of these genes has been explored only in vitro, since the glxR deletion mutant exhibits severe growth defect.In this study, the expression of ptsG and adhA was analyzed in detail indirectly using an adenylate cyclase gene (cyaB) deletion mutant instead of the glxR deletion mutant. The specific activity of ADH was increased in cyaB mutant in glucose and glucose plus ethanol medium, in contrast to the wild-type. In accordance, the promoter activity of adhA was shownderepressed in the mutant, indicating that glxR acts as a repressor of adhA.Similarly, the cyaB mutant exhibited derepression of ptsG regardless of the carbon source. These results confirm the involvement of GlxR, on the expression of important carbon metabolic genes, ptsG and adhA. In addition, we have analyzed the expression of glxR in medium containingvarious carbon sources. Keywords : Corynebacterium glutamicum, GlxR, Adenylate cyclase gene

(cyaB) deletion mutant

E-8

Characterization of a Bacterial Strain Showing High-motility on Solid State Agar Plate Sungwan KANG, Ah Young YOO, Jong Earn YU and Ho Young KANG*Dept. of Lifescience, Pusan National University, Busan 609-735, Korea. *Corresponding author: [email protected]

A bacterial strain running fast on LB media containing 1.5% agar hasbeen isolated from environment. It was moved by extraordinary manor compared with other general motile bacteria. The strain grew well in LB media, however its growth and motility were defected in the presence of several sugars including glucose and maltose. Interestingly, the bacterialstrain showed co-migration when it was mixed with motile Salmonella typhimurium under glucose existing condition. The strain appeared to rod-shaped bacteria under microscopic analyses. In a preliminary study, its morphology was changed to a long-rod shape when it contacts with other motile bacteria. The local area around colonies on agar plate have been observed to form unusual agar shrinking. The strain grew a minimalmedia containing pure agar as a sole carbon source, indicating it may poses agar-degrading activity. In an in-vitro experiment we demonstrated that the cell-free culture supernatant contains agar-degrading activities. Weare now in the middle of typing the strain for classification. Keywords : agar-degrading activity, high-motility, co-migration

E-9

F_ Bioactive Compounds and Chemical Biology

New Phenazine and Pyrrole from a Marine Actinomycete, Strain CNQ-509 JINSOO PARK2, Hak Cheol KWON2 and William FENICAL*1

1Center for Marine Biotechnology and Biomedicine, Scripps Institution of Oceanography, UCSD, La Jolla, USA. 2Natural Products Research Center, KIST,Gangneung 210-340, Korea. *Corresponding author: [email protected]

In the course of our screening for bioactive new secondary metabolitesfrom marine actinomycetes, a new phenzine (1) and a new pyrrole (2), together with previously reported terpenoid naphtoquinone, debromomarinone (3), were isolated from a cultured Streptomyces related bacterium, strain CNQ-509. This strain was isolated from marine sedimentcollected at a depth of 44m offshore of La Jolla, California. The HPLC analysis of culture extract showed that the production of several notable secondary metabolites having unique UV absorbance. These peaks were isolated and purified with repeated column chromatography to yield two new compounds. The structure of new phenazine (1) was determined as 2-linalyl-1,6-phenazinediol-5-oxide and it’s linalyl functionality was presumed to be post-modified by Claisen rearrangement of geranyl group. The structure of new pyrrole (2) is composed of a-carbomethoxypyrrole with farnesyl group at C-4 position, which is the first report of farnesylated pyrrole compound in nature. Based on its production of compound 1, 2, and 3, we can suppose that the strain CNQ-509 has more than two differenct prenyltransferase (PT) genes in its genome or its PT has relaxed substrate-specificity. Keywords : terpenoid, Actinomycete, secondary metabolite

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Isolation of Tyrosinase Inhibition Activity Compound from Biotransformed Root of Elder Platycodin Grandiflorum by Viscozyme Dong-min KANG1, Sun Nyoung YU1, Kwang Youn KIM1, Young Rang JIN1, Hyun Joo JEON1, Ro Ui KIM2 and Soon Cheol AHN*1

1Department of Microbiology and Immunology, Pusan National University Schoolof Medicine, Beomeo-ri, Mulgum-eop, Yangsan 626-770, Gyeoungnam, Korea.2Department of Food Science & Technology, Pusan National University, 50, Cheonghak-ri, Samnangjin-eup, Miryang 627-706, Gyeongnam, Korea. *Corresponding author: [email protected]

Melanin plays an important role in preventing ultraviolet (UV) light-induced skin damage. However, overexposure to UV radiation can lead to a pathologicalincrease in melanin production. Because the tyrosinase is a key enzyme in melanogenesis, the inhibitors of tyrosinase might suppress melanogenesis in the epidermal layer of the skin.The Platycodin grandiflorumA. DC (Campanulaceae) is a traditional food and oriental medicine in Korea, which have been used to treat bronchitis, asthma, pulmonary tuberculosis, diabetesand inflammatory diseases. In this study, we biotransformed P. grandiflorum (22 years old) by enzymes or probiotic microbes and confirmed the enhancedbioactivities in solvents extract fraction. Especially, the highest bioactivities were seen in ethylacetate layer extracts. Moreover, we confirmed a high tyrosinaseinhibition activity in biotransformant by viscozyme. To found the tyrosinase inhibition compounds, extract of biotransformed P. grandiflorum by viscozyme was carried out based on activity-guided fractionation to use column chromatography such as silica-gel, Sepadex LH-20 and HPLC (High Performance Liquid Chromatography).Taken together, we found and isolated the major compound that have the tyrosinase inhibition activity in biotransformed P. grandiflorum. Keywords : Tyrosinase inhibitor, Platycodin grandiflorum, Biotransformation

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Biosynthetic Gene Cluster Engineering for Novel TautomycetinAnalogs Using Streptomyces Targeted Gene Disruption Si Sun CHOI1, David H. SHERMAN2 and Eung-soo KIM*1

1Department of Biological Engineering, Inha University, Incheon 402-751, Korea.2Life Sciences Institute and Department of Medicinal Chemistry, University of Michigan, Ann Arbor, MI 48109, USA. *Corresponding author: [email protected]

Tautomycetin (TMC), which is produced by Streptomyces sp. CK4412, is a novel activated T cell-specific immunosuppressive and anti-cancer compound with an ester bond linkage between a terminal cyclic anhydridemoiety and a linear polyketide chain bearing an unusual terminal alkene.Previously, we cloned, sequenced, and characterized the entire biosynthetic gene cluster for TMC. Sequence information analysis of an 110kb DNAregion revealed multi-modular type I polyketide synthases, type II thioesterase, various proteins for dialkylmaleic anhydride biosynthesis, regulatory proteins and several tailoring enzymes. Although both TMC and its close structural relative called Tautomycin (TM) compounds havesimilar inhibitory activities against two major serine/threonine protein phosphatise, PP1 and PP2A, the different linear polyketide moiety present in TMC and TM is believe to play a critical role because only TMC exhibits an immunosuppressant activity as well as a higher PP1 selectivity.To increase our understanding about the functional specificity, the targetedgene inactivation of various biosynthetic genes involved in the formationof a linear TMC polyketide moiety was performed, leading to the generation of several novel TMC analogs with different polyketide terminalregion. Keywords : Streptomyces, Tautomycetin, immunosuppressant

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Discovery and Biological Activities of New Anti-angiogenic SmallMolecule Ki hyun KIM, Ju Yeol PARK, Hye Jin JUNG and Ho Jeong KWON*Dept. of Biotechnology, Yonsei University, Seoul 120-749, Korea. *Corresponding author: [email protected]

Hypoxia Inducing Factor (HIF)1α plays a key role in the adaptation of tumor cells to hypoxia by activating the transcription of genes, whichregulate several biological processes including angiogenesis. Accordingly,discovering new small molecules that regulate HIF-1α stability is the powerful way to control angiogenesis. First, we selected compounds fromthe well-characterized focused chemical library in a structure-based screen.These compounds were subjected to western blot assay that affected the HIF-1α protein stability. As the result, G0811, was identified as the most potent inhibitor of HIF-1α stability. The compound was treated on HUVECs to confirm the anti-proliferation activity and toxicity. Tube formation, chemoinvasion, and CAM assay were conducted to investigatethe anti-angiogenic activities of the compound both in vitro and in vivo.Furthermore, the effect of the compound on reactive oxygen species (ROS), a key player in angiogenesis, was investigated to elucidate the mechanism underlying for anti-angiogenic activity. G0811 effectively decreased DCF fluorescence intensity, demonstrating that the compound decreases ROS levels in the cells. Collectively, we were able to discover new anti-angiogenic small molecule targeting ROS-mediated pathway in the cells. Keywords : Hypoxia Inducing Factor-1α, angiogenesis, small molecule

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Production and Characterization of Siderophore from Pseudomonas sp. Tur-4 under Chitosanase Production ConditionKyoung-Ja KIM*, Yong JIN and Gwang-hyun LEEDept. of Biotechnology, University of Soonchunhyang, Asan City, Chungnam, 336-745 Korea. *Corresponding author: [email protected]

Carboxylate type siderophore was produced from Pseudomonas sp. Tur-4 under chitosanase production condition. Siderophore production was higher at the early exponential phase than that at the late exponential phaseof growth. But the chitosanase production was higher at the late exponentialphase of growth than that at the early exponential phase. A decrease in siderophore production was observed with an increase in iron concentration. 20 M FeCl3 inhibited the production of siderophore completely, but the production of chitosanase was not inhibited by 20 M FeCl3. Partially purified outer membrane receptor protein for siderophore under iron deficient condition was identified on the SDS-PAGE. Nitric oxide binding activity and antimicrobial activity of siderophore were determined. Exogeneously added siderophore into the culture medium of Streptomyces lincolnensis M-20 acts as a inducer for siderophore production.Keywords : siderophore, chitosanase, outer membrane receptor, nitric oxide

binding activity, antimicrobial activity.

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Isolation of Diketopiperazine and Related Metabolites from the Marine-derived Fungus Penicillium sp. SF-5203 Yu-Ri LEE1, Su-Bin SOHN1, Chi-Wook LEE1, Changon SEO1, Jae-Hak SOHN1, Jong Seog AHN2 and Hyuncheol OH*1

1Dept. of Bio-Food Materials, Silla University, Busan 617-736, Korea. 2Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 305-333, Korea. *Corresponding author: [email protected]

In the course of the screening program for PTP1B inhibitors from marinemicrobial resources, a marine-derived fungal isolate SF-5203 was selected for further investigation on the basis of its potent inhibitory effect against PTP1B. The marine-derived fungus Penicillium sp. SF-5203 was isolatedfrom the shoreline sediment sample collected at Wan Island. The methyl ethyl ketone extract of the fungal culture was subjected to C18 functionalized silica gel flash column chromatography, eluting with a stepwise gradient consisting of MeOH in H2O. The fraction eluted at 60%MeOH was then subjected to semi-preparative reversed-phase HPLC to yield compounds 1 and 2. Compound 3 was isolated from the fraction eluted at 70% MeOH. Compounds 1-3 were identified as fructigenine A(1), cyclopenin (2), and cyclopenol (3) mainly by analysis of NMR and MS data. Fructigenine A has been reported to have inhibitory activity against oat coleoptiles and L-5178y cells. In this study, fructigenine A (1) was shown to inhibit the activity of PTP1B with IC50 value of 10 μM, and the inhibition mode was evaluated by kinetic study. Details of the isolation, structure determination, and their PTP1B inhibitory activitieswill be presented Keywords : marine fungus (Penicillium sp.), Diketopiperazine, PTP1B

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New Biological Function and Application of Low Molecular Weight Poly-γ-glutamic Acid Chung PARK1, Haruyoshi SEINO3, Kazuhiko HAMADA3, Mie IWAMOTO1, Hiroshi UYAMA4, Chul-Joong KIM5, Haryoung POO6 and Moon-Hee SUNG*1,2

1BioLeaders Corporation, Daejeon 305-500, Korea. 2Department of Advanced Fermentation Fusion Science and Technology, Kookmin University, Seoul 136-702, Korea. 3Pias Corporation, Osaka 531-0072, Japan. 4Department of Applied Chemistry, Osaka University, Suita 565-0871, Japan. 5Department of Veterinary Medicine, Chungnam National University, Daejeon. 6Pandemic Influenza Preparedness Research Center, KRIBB, Daejeon 305-333, Korea. *Corresponding author: [email protected]

Poly-γ-glutamic acid (γ-PGA) is a most promising natural biopolymerthat is produced by Bacillus subtilis (chungkookjang). Various qualities of γ-PGA include the ability to form a smooth, elastic, self-moisturizingand soft film on the skin, resulting improved sensory perception and protection of the outer layer of skin. At present, we succeeded in producingγ-PGA with low molecular weight γ-PGA industrially and developing newfunctional applications in functional cosmetics. The results showed that 1,000 Da γ-PGA penetrate skin and increase hyaluronic acid in the basal layer in skin and suggested that γ-PGA reinforce the skin’s support structure to energize cells, renewal of the skin. The low molecular weight γ-PGA may be very useful in the development of new anti-aging cosmetics. [This work was supported by a Seoul R&BD program (10580)’ in Korea program grant from the Seoul Development Institute.] Keywords : low molecular weight poly-γ-glutamic acid, anti-aging effect,

functional cosmetics

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Hypholomine B and Hispidin, Inhibitors of enoyl-ACP Reductase Produced by the Fungus Phellinus Linteus Yun-ju KWON1, Jun-Young CHO1, Mi-Jin SOHN1, Soon-Ja SEOK2 and Won-Gon KIM*1

1Korea Research Institute of Bioscience and Biotechnology, P. O. Box 115, Yusong, Daejeon 305-806, Korea. 2National Institute of Agricultural Science andTechnology, RDA, Suwan 441-707, Korea. *Corresponding author: [email protected]

Bacterial enoyl-ACP reductase (FabI) that catalyzes the final and rate-limiting step in the type II fatty acid synthesis has been validated as a novel target for antibacterial drug development. The medicinal fungusPhellinus linteus in the family Hymenochaetaceae has been used as a traditional medicine for the treatment of various diseases. In the course of our screening for FabI inhibitors, the hispidin dimer hypholomine B and hispidin monomer were isolated from a culture broth of a medicinal fungus Phellinus linteus. Their chemical structures were identified by MS and NMR spectral analysis. Hypholomine B and hispidin inhibited Staphylococcus aureus enoyl-ACP reductase with IC50 of 4 -5 uM, while caffeic acid didn’t inhibited even at 100 uM.. They showed 16 and 40 times higher inhibition against S.aureus FabI than Streptococcus pneumonia FabK and Mycobacterium tuberculosis InhA, respectively. Thehispidin also showed antibacterial activity against Staphylococcus aureus, Bacillus subtilis, and MRSA. This work was supported by the 21C Frontier Microbial Genomics and Application Center Program. Keywords : Phellinus linteus, Hypholomine B, hispidin

F-8

Isoflavone Contents and Electrophoresis Pattern of Chunggukjang Prepared with Various Soybean Cultivars So Young KIM1, Min Hwa KIM1, Jong Min KO2 and Yong Suk KIM*1

1Department of Food Science & Technology, Chonbuk National University, Jeonju, Jeonbuk 561-756, Korea. 2Department of Functional Crop, National Institute of Crop Science, RDA, Korea. *Corresponding author: [email protected]

To select the optimal soybean cultivars for Chunggukjang preparation, isoflavone contents and electrophoresis pattern were compared in 30 soybean cultivars. Isoflavone has antioxidant, anticancer activity and estrogen-like activity of functional ingredients. Chunggukjang prepared with various soybean cultivars showed significant difference in total isoflavone contents. Total isoflavone contents of Chunggukjang prepared with Daepung were the highest 208.75 mg% and Nampung, Sodam, Pungsannamul, and Wonkwang were 187.11 mg%, 149.68 mg%, 147.11 mg%, 143.41 mg%, respectively. Among the Chunggukjang of series in black, Cheongja#3 showed the highest 120.15 mg%. SDS-PAGE Electrophoresis pattern results of Chunggukjang prepared with various soybean cultivars showed mostly a broad band in the range of 26 kDa. Especially, 26kDa band in the Cheongja, Cheongja#2, Cheongja#3, Heukmi, Daeheuk, and Heukseong Chunggukjang could be seen appeared bold. Electrophoresis pattern of Chunggkjang prepared with various soybean cultivars was confirmed mostly to form low molecular weight peptide. From these results, the functional effects like ACE inhibition activity or antioxidant activity of Chunggkjang were confirmed indirectly.

F-9

Purification of Determinants of Ochrobactrum Lupini KUDC1013 Inducing Systemic Resistance Against a Soft-Rot Pathogen in Tobacco Marilyn SUMAYO1, Mi-Seon HAHM1, Ye-Ji HWANG1, Choong-Min RYU2

and Sa-Youl GHIM*1

1School of Life Sciences and Research Institute for Ulleungdo and Dokdo, Kyungpook National University and Research Institute for Ulleungdo and Dokdoand School of Life Sciences, Kyungpook National University, Daegu 702-701, Korea. 2Laboratory of Microbial Genomics, Systems Microbiology Research Center, Korea Research Institute of Bioscience and Biotechnology, P.O. Box115,Yuseong Daejeon, Korea. *Corresponding author: [email protected]

Ochrobactrum lupini KUDC1013 is a plant growth promoting rhizobacterium (PGPR) isolated from the rhizosphere of Solanum nigrumL. in Dokdo island. It has the capability to induce systemic resistance (ISR) of tobacco against the leaf soft-rot pathogen Erwinia carotovora subsp. carotovora strain SCC1. KUDC1013 produces siderophore and has flagella and LPS, compounds that are known to induce systemic resistance.However, the mechanism and metabolites of KUDC1013 eliciting ISR activity remain unknown. To identify and purify compounds that provideinduced systemic resistance, extracellular components produced by KUDC1013 were extracted with series of solvents, fractionated, purified and were tested for their protective effects. ISR bioassay revealed that resistance elicitors were retained in the cell free supernatant and was partitioned into n-butanol. KUDC1013 is a potential biocontrol agent. Further purification and identification analyses of the ISR elicitor from KUDC1013 will increase the range of factors from PGPR recognized to induce plant resistance.

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Morus alba L. Root Bark Extract Stimulates Adipose Differentiation in 3T3-L1 Cells by Up-regulating Adipogenic Transcription Factors and Genes Taeseok OH, Duk Kwon CHOI and Jong Won YUN*Dept. of Biotechnology, Daegu University, Kyungsan, Kyungbuk 712-714, Korea.*Corresponding author: [email protected]

The root bark of mulberry tree (Morus alba L.) has been traditionally used in Asian countries for medicinal purposes due to its anti-inflammatory, hypoglycemic, antibacterial, and antiviral effects. In the present study, it was found that the Morus alba L. root bark extract (MARE) stimulated 3T3-L1 preadipocytes differentiation. Treatment with MARE enhanced the triglyceride (TG) content of the cultures and increased the levels of C/EBPα(CCAAT/enhancer-binding protein α) and PPARγ (peroxisome proliferator-activated receptors γ) mRNAs during adipocyte differentiation.Immunoblot analysis indicated that MARE increased the expression of adipogenic makers such as PPARγ, adipocyte-specific fatty acid bindingprotein (aP2), and GLUT4 (glucose transporter 4). Taken together, our results suggest that MARE promoted adipogenesis in 3T3-L1 preadipocytecells through PPARγ-related pathways at both transcript and protein levels.Although these results require further studies, this natural plant extract may be expected to be used as a possible therapeutic material for dyslipidemia, type 2 diabetes mellitus, and postmenopausal metabolic syndrome with hypoadiponectinemia by increasing PPARγ transcriptionalactivities. Keywords : Morus alba L., 3T3-L1 preadipocyte, Adipogenesis

F-11

A New Derivative of Aquastatin A from Sporothrix sp. FN611 Yun-ju KWON, Mi-Jin SOHN, Tae-Kwon OH and Won-Gon KIM*Korea Research Institute of Bioscience & Biotechnology, Yusong, Daejeon 305-806, Korea. *Corresponding author: [email protected]

Previously we reported that aquastatin A was isolated as a FabI inhibitorfrom liquid fermentation cultures of Sporothrix sp. FN611. Further investigation of minor and less polar metabolites of Sporothrix sp. AT 28 have resulted in isolation of a new derivative of aquastatin A. Its chemical structure was determined by MS and NMR spectral data. The new compound doesn’t contain a 2-hydroxy-6-methyl benzoic acid moietyof aquastatin A. Interestingly, it lost inhibitory activity against bacterial enoyl-acyl carrier protein (ACP) reductases including FabI, FabK, and InhA, and also antibacterial activity of aquastatin A. These results show that the 2-hydroxy-6-methyl benzoic acid moiety is critical for inhibitoryactivity of aquastatin A against enoyl-ACP reductases and bacterial growth. This work was supported by the 21C Frontier Microbial Genomics and Application Center Program. Keywords : Aquastatin, enoyl-ACP reductase, antibacterial

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Effect of Antimicrobial Compounds with Marine Actinomycete,Streptomyces Microflavus Strain CY-2 on Immune Response of Olive Flounder, Paralichthys OlivaceusIk Soo JANG2, Man-chul KIM2, Li-Na KIM1, Ju-Sang KIM2 and Moon-soo HEO*1

1Dept. Marine Biomedical Sciences, jeju-University, Jeju 690-120, Korea. 2Dept. Marine life Science, jeju-University, Jeju 690-120, Korea. *Corresponding author: [email protected]

Streptomyces microflavus strain CY-2 of marine actinomycetes were obtained from Jeju Island. The effects of antimicrobial compounds produceby marine actinomycetes Streptomyces microflavus strain CY-2 cultural extract supplemented diet on hematology and disease resistance in olive flounder, Paralichthys olivaceus against Streptococcus iniae. Growth rateof Paralichthus olivaceus was found to be 154.4 g group fed with normal diet. However it was found to be 174.3 g in group feeding material withof SBME-18 of 1%. Blood GOT of experimental fish did not significantdifference after comparing with control. Blood GTP enzyme of experimental fish had no significant difference compared to control. Oxygen free radical in phagocyte of fish, was found to be higer in groupfed with supplemented diet group. Mortality rate of Paralichthus olivaceus low in group fed with actinomycetes extract diet after 13 weeks against Streptococcus iniae. Keywords : Antimicrobial Compounds, Marine Actinomycete, Streptomyces

microflavus

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Biotransformation Enhances Rhapontin Antimicrobial Activity and Provides Clues to its Mechanism of Action Young Hee LIM*1, Narae KIM1, Jeong-Keun KIM2, Hyun-Sun LEE3, Keun-Tae PARK1

1Department of Biomedical Science, College of Health Science, Korea University.2Department of Chemical Engineering and Biotechnology, Korea Polytechnic University. 3Department of Food and Nutrition, College of Health Science, KoreaUniversity. *Corresponding author: [email protected]

Biotransformation is often used to improve biological activity of prodrugs. Rhapontin was isolated and identified from the methanol extract of the roots of Rheum undulatum. Rhapontigenin, the aglycone of rhapontin, was biotransformated from rhapontin by Pectinex®, and evaluated the in vitro antimicrobial activity. Rhapontigenin showed 4- to16-fold higher antimicrobial activity than did rhapontin. The antimicrobial activity of rhapontigenin was higher against Gram positive and fungal strains than Gram negative strains. Moreover, cytotoxic activityof rhapontigenin was not observed in hemolytic assay within the concentration used for the antimicrobial activity assay. Rhapontigenin mainly inhibited protein synthesis and RNA synthesis in Staphylococcus aureus and Candida albicans, respectively. Rhapontigenin induced morphological changes of S. aureus and C. albicans, which might have contributed to cellular growth inhibition. In conclusion, the biotransformation of rhapontin into rhapontigenin conferred antimicrobialactivity against S. aureus and C. albicans at concentrations that were notcytotoxic to eukaryotic cells.Keywords : Biotransformation, Rhapontigenin, Antimicrobial activity

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Arenicin-1 Triggers Apoptosis in Candida Albicans Jaeyong CHO, Juneyoung LEE and Dong Gun LEE*School of Life Sciences and Biotechnology, College of Natural Sciences, Kyungpook National University, Daegu 702-701, Korea. *Corresponding author: [email protected]

Arenicin-1, a 21-residue cationic peptide isolated from marine polychaeta Arenicola marina, is known to contain antimicrobial activities.In our previously study, we investigated Arenicin-1 had an antifungal effects through membrane-active mechanism. To understand the further mode of action(s) of Arenicin-1 after the pertubing fungal membrane, weconducted experiments on Candida albicans which is human infectious fungi as causing candidiasis. The result showed that Arenicin-1 bound to DNA and affected the cell cycle progress of C. albicans. Flow cytometricanalysis, using DiBAC4(3), DiOC6(3), DHR-123 and Annexin-V-FITC, showed that Arenicin-1 induced plasma membrane depolarization, mitochondrial membrane depolarization, production of reactive oxygen species (ROS) and phosphatidylserine externalization which are commom hallmarks of apoptosis. Furthermore, TUNEL assay and DAPI staining assay indicated that Arenicin-1 caused DNA fragmentation and condensation in C. albicans. Therefore, these results suggest that Arenicin-1 exerts an antifungal effect against C. albicans by inducing apoptosis. Keywords : Apoptosis, Arenicin-1, Antimicrobial peptide

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Preparations and Characteristics of Poly-γ-Glutamic Acid Hydrogels Su-jung HAM1, Chung PARK2, Ok Kyoung JEON1, Doo-Yeol CHOI1, Mi-Sun KWAK1, Iwamoto MIE3, Uyama HIROSHI4 and Moon-Hee SUNG*1,2

1Dept. of Advanced Fermentation Fusion Science and Technology, Kookmin University, Seoul, Korea. 2BioLeaders Corporation, Daejeon, Korea. 3BioLeadersCorporation, Osaka, Japan. 4Graduate School of Engineering, Osaka University, Suita, Japan. *Corresponding author: [email protected]

Naturally occurring polymers have attracted considerable interest among polymer scientists in recent years. Poly-γ-glutamic acid (γ-PGA) is naturally synthesized by Bacillus subtilis (chungkookjang), and is biodegradable. γ-PGA hydrogel can be used as a compatible biomaterial for many applications. We prepared several types of hydrogel using γ-PGA. γ-PGA hydrogel by γ-irradiation showed individual specific water content and fast swelling property. This material has cross-linked networks and pore structures of various sizes. It is possible to regulate the physical property of a hydrogel by the degree of γ-irradiation and the molecular weight of γ-PGA. Also, we prepared a chemical cross-linked gel and a physical gel by using γ-PGA. Each of these gels has different physical characteristics. γ-PGA hydrogel may be used as a functional cosmetics material, a biomedical material, and so on. This indicates that γ-PGA hydrogel could be used in the various fields of industrial application. [This work was supported by 'Seoul R&BD Program (10580)] Keywords : PGA, Hydrogel, Bacillus subtilis

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Antibacterial, Antiviral, and Antioxidative Activities in Thailand Plant Extracts Soo-Ki KIM*1, Joon-Hyeong LEE1, Sang-Beum CHO1, Sang -Heui SEO2 andHyun-Dong PAIK1

1College of Animal Bioscience and Biotechnolgy, Konkuk University, Seoul 143-701, Korea. 2College of Veterinary Medicine, Chungnam National University, Daejeon 305-764, Korea. *Corresponding author: [email protected]

Tropical plant extracts from Thailand were screened for the developmentof livestock feed additives. Forty-two solvent fractions from seven plantswere tested against animal pathogenic bacteria (Escherichia coli, Burkholderia sp, Clostridium perfringens, Pantoea agglomerans, Haemopillsomnus, and Haemopill parsuis) by using paper disk methods. Methanol extracts of Makrut showed antibacterial activity to all tested strains exceptfor Haemopill somnus. Water and methanol fractions of Mark showed the activity specifically to Burkholderia sp. Only butanol fraction from Makkhuayao extracts showed the antibacterial activity for Pantoea agglomerans. Among the tested plants, Keurachaita was found to be a potent inhibitor of H5N1 influenza virus in MDCK cell culture. Butanol extracts of Keurachaita showed the virus titers of 2 log whereas control was 7 log, indicating that Keurachaita inhibited the infection of H5N1 influenza viruses. Antioxidative activity was tested for free radical scavenging effect of DPPH (1,1-diphenyl-2-picrylhydrazyl radical). High antioxidant activities were detected in methanol fractions of Makrut and Mak, chloroform fractions of Khing, butanol fraction of Makkhuayao andMakkhuapro. (This work was supported by Biogreen 21) Keywords : Plant extracts, Antimicrobial activity, Antioxidative activity

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The Effect of Mycelial Culture Broth of Mushrooms on the Expression of Wrinkle care and Moisturizing Related Genes.Yun Jeong KIM*, Jong Woo CHEON and Hyun Sang LEER&D center, ACT Co., Ltd. *Corresponding author: [email protected]

To develop wrinkle care and moisturizing cosmetic ingredient with mushrooms, mycelium of 5 mushroom species were investigated. RT-PCRand Zymography were conducted to identify the efficacy of mycelia culturebroth on the expression of procollagen (COL1A1) and hyaluronan synthase(HAS) genes. Accordingly, Cordyceps militaris, Sparassis crispa and Monascus pilosus were selected as candidates for developing cosmetic ingredient. The effect of carbone and nitrogen sources for the expression of the genes were investigated. Maltose as a carbone source and peptoneas a nitrogen source based on the CMM medium were the optimal carboneand nitrogen for the expression of the genes. The optimal temperature and pH were 30 ℃ and 6.0, respectively. Therefore mycelial culture brothof tested mushrooms could be a useful cosmetic ingredient as a wrinkle care and skin moisturizing. Keywords : wrinkle care, moisturizing, mushroom

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A Functional Study on Physiologically Activating Compounds Produced from Purple Nonsulfur Bacteria Through Carbon Dioxide Fixation Hyun Jeong LEE1, Ju-Yong PARK1, Thai-Hoang LE2, Yang-Hoon KIM3 and Jiho MIN*1

1Graduate School of Semiconductor and Chemical Engineering, Chonbuk National University, 664-14 Duckjin-dong, Jeonju 561-756, South Korea. 2Department of Bioprocess Engineering, Chonbuk National University, 664-14 Duckjin-dong, Jeonju 561-756, South Korea. 3School of Life Science, Chungbuk National University, 410 Sungbong-Ro, Heungduk-Gu, Cheongju 361-763, South Korea. *Corresponding author: [email protected]

In Purple nonsulfur photosynthetic bacteria, carbon dioxide fixation has been intensively investigated, and through the Calvin reductive pentose phosphate pathway by ribulose 1,5-bisphosphate carboxylase/oxygenase, this organism can produce organic compounds such as sugar from carbon dioxide. This study is focused on the impact on physiological activation of other microorganisms by carbon dioxide fixed organic compounds from Rhodobacter sphaeroides cultured differential light condition (PACs). Wehave constructed recombinant Escherichia coli harboring hupSL hydrogenaseisolated from R. sphaeroides, and we also found that biohydrogen production from recombinant E. coli was increased under the PACs supplemented condition. In comparison with NCS, dark PACs have high efficiency for mammalian cell growth. In addition, we found that baby daphnia numberwas increased by adding the dark PACs in Daphnia magna. Analysis of the bacteriochlorophylla and carotenoid pigment extracted from cells of dark condition shows a increase in comparison with light and light:dark condition, which correlates with increase in the physiological activation. From these results, we conclude that PACs was not only the carbon source,but also nutrient and physiological activation on the growth of microorganism. Keywords : Rhodobscter sphaeroides, dark PACs, physiological activation

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Label-Free Detection with Polydiacetylene Biochip Using a NovelImmobilizing Method Moo-Kyung PARK, Kyung-Woo KIM, Dong June AHN and Min-Kyu OH*Dept. of Chemical and Biological Engineering, Korea University, Seoul 136-713, Korea. *Corresponding author: [email protected]

In order to detect sequence-specific single stranded DNA (ssDNA), a new type of immobilization method was developed in the design of a polydiacetylene (PDA) liposome-based label-free biochip. A few PDA immobilization methods have been introduced, but most of them had drawbacks in availability or efficiency. To overcome these problems, we utilized the α-cyclodextrin (CD) that strongly binds to the terminal structure of PDA liposomes. PDA liposomes were immobilized on α-CD coated substrates. By using this strategy, we were able to immobilize them moreefficiently and rapidly. The structures of immobilized PDA were characterized by scanning electron microscopy (SEM). Target ssDNAs were successfully detected by rapid hybridization (<1hr) to its complementary DNA probe on PDA liposome. The fabricated biochip was able to perform the specific and quantitative detection of ssDNA. The result of this study showed that an advanced DNA detection method by label-free biochip was successfully developed. Keywords : Label-free biochip, Polydiacetylene, DNA hybridization

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Evaluation of the Inhibition of Melanin Synthesis of the Extractfrom Morus alba by Biotransformation Young Hee LIM*1, Jeong-Keun KIM2, Hyun-Sun LEE3 and Mijin KIM2

1Departmnet of Biomedical Science, College of Health Science, Korea University.2Department of Chemical Engineering and Biotechnology, Korea Polytechnic University. 3Department of Food and Nutrition, College of Health Science, KoreaUniversity. *Corresponding author: [email protected]

Enzymatic conversion of glucosides into aglycones improves their biological activities. Mulberroside A, a glycosylated stilbene, was isolated and identified from the ethanol extract of the roots of Morus alba. Oxyresveratrol, the aglycone of mulberroside A, was produced by enzymatic hydrolysis of mulberroside A using the commercial enzyme Pectinex®. Mulberroside A and oxyresveratrol showed inhibitory activityagainst mushroom tyrosinase, with oxyresveratrol demonstrating a greaterenhanced inhibitory effect than mulberroside A on mushroom tyrosinase activity. Oxyresveratrol and mulberroside A strongly inhibited melanin production by Streptomyces bikiniensis, and manifested dose-dependent tyrosinase inhibiting activity and inhibition of melanin synthesis in B16F10 melanoma cells. However, they exhibited nearly similar inhibitory effectson the activity of cellular tyrosinase and melanin synthesis in melanocytes.Oxyresveratrol and mulberroside A displayed a UV-blocking effect at UVAand UVB ranges. These results indicate that oxyresveratrol produced by biotransformation from mulberroside A may be suitable for use as a potentskin-whitening agent. Keywords : Biotransformation, Melanin synthesis inhibition, Oxyresveratrol

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Effects of Extracts and Fractions from Peel and Seed of Avocado on Osteoblast Differentiation and Osteoclast Formation Mi-Jin KIM2, Nam-Kyung IM2, Hyuk-Il KIM2, In-Seon LEE2 and Hyun-Jeong KIM*1

1The Center for Traditional Microorganism Resources, Keimyung University.2Department of Food and Technology, Keimyung University. *Corresponding author: [email protected]

The avocado (Persea americana Mill.) is a widely grown and consumed fruit. It has been known to have some biological activities such as acetylCoA carboxylase inhibition, antioxidant and antimicrobial activity. In general, bone is continuously remodeled by osteoblasts and osteoclasts. To investigate the bioactivity of avocado which acts on bone formation, we searched the effects of extracts and fractions from peel and seed of avocado on the activities of osteoblast and osteoclast cells. The activity in bone formation with MC3T3-E1 cells and bone resorption with osteoclasts, derived from mouse bone marrow macrophages, were measured. The methanol extracts, hexane fr. and ethylacetate fr. from peel of avocado showed stronger bone-forming activity accompanied with osteoblasts proliferation and alkaline phosphatase (ALP) activity than avocado seed extracts. In addition, avocado peel extracts also decreased tartrate-resistant acid phosphatase (TRAP) activity against differentiation of osteoclasts. Therefore, avocado is expected to be a natural sauce for the developments of medicinal agents to prevent bone-related diseases suchas osteoporosis by increasing the differentiation of osteoblasts and reducingthe activity of osteoclasts. Keywords : Abocado, Osteoblast, Osteoclast

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Anti-Allergic Effect of Exo-Biopolymer in NC/Nga Mice Modelof Atopic Dermatitis Seonggil HONG, Joong-Hark KIM, Hwa-young KIM, Sung-Joo HWANG and Dong Ju KIM*Erom R&D center, Erom Co. Ltd, Chungcheon, Korea. *Corresponding author: [email protected]

The purpose of this study was to evaluate the anti-allergic effects of a substance extracted from a submerged culture of Lentinus edodes withrice bran (RBF) and secondary fermentation of RBF with Lactobacilus (RBFLB) in the NC/Nga mice. The NC/Nga mice were induced atopic dermatitis using by dinitrochlorobenzene(DNCB) for 2 weeks. The mice were divided into five groups: normal control (NC), control (C), RBF, RBFLB and biobran (BR) as similar product of RBF. Each sample was administrated orally to the NC/Nga mice during 4 weeks of experimentalperiod. Result of the experiment, the modified SCORAD score was significantly decreased in the RBF, RBFLB and BR group compared to the control group. The levels of cytokine (IL-4, IL-10) released from spleenocyte were reduced and the level of IFN-r was significantly increasedcompared to the control group. In addition, the IgE concentration of plasmaand splenocyte were significantly decreased compared to the control group. In the results of this study, we suggest that RBF, RBFLB and RB have potentially useful anti-allergic effects mediated by regulation of cytokinesand IgE levels. Keywords : rice bran, fermentation, NC/Nga mice

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Purification of Antibacterial Compounds Produced by Streptomyces sp. MJM6010Dong shik KIM1, In Ae LEE2 and Joo won SUH*1

1Division of Bioscience and Bioinformatics, Myongji University, Yongin, Gyeonggi-Do, Korea. 2Division of Bioscience and Bioinformatics, Institute of Natureal Science, Myongji University, Yongin, Gyeonggi-Do, Korea. *Corresponding author: [email protected]

We screened actinomycetes which produce potent antibacterial compounds from ECUM(Extract Collection of Useful Microorganism) inMyongji univ. and identified as Streptomyces sp. MJM6010. We purified the antibacterial compounds from 10-liter culture broth by Ethyl acetate solvent, RP-18 reversed vacuum flash column chromatography, and RP-18preparative HPLC with 45% aqueous acetonitrile and 63% aqueous Methanol. The structure of purified antibacterial compounds were Quinomycin derivatives by 1H-NMR and mass data. We will determine the exact structures by 2D-NMR. The antibacterial compounds from MJM6010 exhibited a strong antibacterial activity against Staphylococcusaureus and Methilcillin-resistant S. aureus with minimal inhibitory concentration (MIC) in the range of 0.2~10 ㎍/㎖. Keywords : Streptomyces sp, Methilcillin-resistant S. aureus,

Staphylococcus aureus

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Effect of Dropwort Extracts Fermented with Oligosaccarides on Allergic Inflammation EUN JU LEE1, Seun-Ah YANG3, Hyung-Dae KIM4, Hyo-Gwon IM5, Sam-Pin LEE1,2 and In-Seon LEE*1,2

1Dep. of Food Science and Technology, Keimyung University, Daegu, Korea. 2TheCenter for Traditional Microorganism Resources, Keimyung University, Daegu, Korea. 3Center for Traditional Microorganism Resources, Keimyung University, Daegu, Korea. 4Bissell Chunglog Farm, Daegu, Korea. 5Daegu Technopark Bio Industry Center, Daegu, Korea. *Corresponding author: [email protected]

Dropwort (Oenanthe javanica) is a medicinal food commonly used to treat diseases such as jaundice, hypertension, and polydipsia in Korea, Japan, and China. In this study, the anti-allergic inflammation effects of dropwort extracts were evaluated using RBL-2H3 mast cells and RAW 264.7 cells. Dropwort extracts were fermented by steeping with oligosaccaride at room temperature for 1 year (1 yr), and then stored at 4°C for 1 (2 yr) or 2 year (3 yr). The extracts (0.01 - 1.0 mg/mL) had no cytotoxicities on IgE-sensitized RBL-2H3 cells, and the 2 yr (25% at 1 mg/mL) and 3 yr (39% at 1 mg/mL) extracts inhibited the beta-hexosaminidase release. However, the 1 yr extract had little effect on the beta-hexosaminidase release. In addition, the inhibitory effects onthe productions of NO and proinflammatory cytokines in LPS-stimulated RAW 264.7 cells were tested to evaluate optimal againg period for anti-inflammatory potential. Keywords : Dropwort, Oenanthe javanica, allergic inflammation

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Antifungal Effects of Psacotheasin Promoting Apoptosis via Membrane-Active Mechanism on Candida albicans Bomi HWANG1, Juneyoung LEE1, Jae-Sam HWANG2 and Dong Gun LEE*1

1School of Life Sciences and Biotechnology, College of Natural Sciences, Kyungpook National University, Daegu 702-701, Korea. 2National Academy of Agricultural Science, Rural Development Administration, Suwon 441-100, Korea. *Corresponding author: [email protected]

Psacotheasin is a 34-mer knottin-type peptide derived from Psacothea hilaris. In this study, we investigated the antifungal effects of psacotheasinand mechanisms in Candida albicans. Psacotheasin showed remarkable antifungal activity, against some kinds of human pathogenic fungi, withouthemolysis on human erythrocytes. To understand the antifungal mechanisms of psacotheasin, the fungal membrane interactions of peptidewere examined. Flow cytometric analysis with DiBAC4(3) and PI exhibitedthat psacotheasin depolarized and disrupted the membrane of C. albicans. These results were confirmed by 3D-flow cytometric contour-plot analysis.The further studies, using FITC-dextran loaded liposome and rhodamine-labeled GUV, indicated that psacotheasin act as pore-formingantifungal peptide in the membranes of C. albicans. Additionally, psacotheasin induced a series of cellular changes characteristic of apoptosisin C. albicans, including accumulation of ROS, externalization of phosphatidylserine at the outer plasma membrane, reduction of mitochondrial transmembrane potential, significant cell cycle arrest, and DNA condensation and fragmentation. Therefore, this study suggests thatpsacotheasin exert its antifungal activities both by disrupting fungal membranes and by promoting apoptosis. Keywords : Psacotheasin, Membrane-active mechanism, Apoptosis

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Anti-Biofilm Activity of Xanthorrhizol against Vancomycin Resistant Enterococci (VRE) BiofilmsYaya RUKAYADI1, Sunghwa HAN2, Dongeun YONG3 and Jae Kwan HWANG*1

1Depart. of Biotechnology, Yonsei University, Seoul 120-749, Korea. 2Dept. of Biomaterials Science and Engineering, Yonsei University, Seoul 120-749, Korea.3Dept. of Laboratory Medicine & Research Institute of Bacterial Resistance, Yonsei University. Seoul 120-749, Korea. *Corresponding author: [email protected]

Enterococci are the most common Gram-positive cocci in the intestinaltract and have been documented to cause infection of the urinary tract and other sites of humans. In addition, increased antimicrobial resistance in enterococci has become a problem because of their ability to form biofilm, facilitating their persistence in the host. The aim of this study was to investigate the anti-biofilm activity of xanthorrhizol against 23 isolates of clinical VRE. The results showed that MICs of xanthorrhizol against 23 clinical VRE isolates range 0.5 - 4 µg/ml, meanwhile sessile minimum inhibition concentration (SMICs) of xanthorrhizol against 23 isolates of clinical VRE was 2-fold - to 16-fold (4 - 16 µg/ml) greater than the concentration required to inhibit planktonic growth (MIC). Xanthorrhizol was able to eradicate the VRE bioflms with biofilm eradication concentration (BEC) range 32 - 64 µg/ml. These results suggestthat xanthorrhizol can be used to inhibit the growth and eradicate the biofilm formed by clinical VRE.Keywords : Anti-Biofilm, Vancomycin resistant enterococci, Xanthorrhizol

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A New Cyclopentenone from an Edible Marine Seaweed-Associated Fungus Rhizopus stolonifer Byeng Wha SON*, Keumja YUN, Viviane N. NENKEP, Zhile FENG, Guohua YANG, Xavier SIWE-NOUNDOU and Alain S. LEUTOUDepartment of Chemistry, Pukyong National University, Busan 608-737, Korea. *Corresponding author: [email protected]

As part of an effort to discover biologically active natural products frommarine sources, we investigated the bioactive secondary metabolites of a marine isolate of the fungus Rhizopus stolonifer, isolated from the surfaceof the edible marine brown alga Sargassum horneri. The cultured mediumof R. stolonifer was filtered through cheesecloth to give mycelium and broth. The filtered broth was extracted with EtOAc to afford broth extract (1.0 g). The broth extract of R. stolonifer was subjected to silica gel flash chromatography. Elution was performed with n-hexane - EtOAc (stepwise, 0 - 100 % EtOAc) to yield four fractions. Fraction 3 on medium pressureliquid chromatography (MPLC) (ODS) by elution with H2O-MeOH (from 1:1 to 1:5) afforded crude botrytinone, which was further purified by HPLC(YMC, ODS-A) utilizing a 30 min gradient program of 50% to 100% MeOH in H2O to furnish botrytinone (6.6 mg). The molecular structuresand absolute stereochemistry of botrytinone was determined from chemical and physicochemical evidence, including quantum chemistry calculations,X-ray analysis, and the circular dichroism (CD) exciton chirality method. Keywords : Cyclopentenone, Marine-derived fungus, Rhizopus stolonifer

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Phellodendri Cortex Inhibits Differentiation of 3T3-L1 Adipocytes through Suppressing Adipogenic Transcription Factors and GenesDuk Kwon CHOI1, Tae Seok OH2 and Jong Won YUN*1

1Department of Biotechnology, Daegu University, Kyungsan, Kyungbuk 712-714, Korea. 2Department of Biotechnology, Daegu University, Kyungsan, Kyungbuk, Kyungbuk 712-714, Korea. *Corresponding author: [email protected]

Abstract Obesity is a medical disorder in which excess body fat has accumulated, and closely associated with differentiation of adipocyte. Therefore, there have been studies on inhibitor of adipocyte differentiation for prevention of obesity. We screened possible anti-obesity materials among natural plant resources and found that Phellodendri cortex (PC), which has used to as transitional therapeutic of anti-inflammation. 3T3-L1 preadipocytes were treated with a berberine-rich extract of PC; its inhibitory effect on adipocyte differentiation was demonstrated by MTT assay, Oil Red O staining, and TG content. In addition, its effects on obesity-related genes and proteins were validated by Western blot and real-time RT-PCR analysis. Consequently, PC significantly inhibited differentiation of 3T3-L1 cells and lowered TG level. Moreover, expressionlevels of PAARg and C/EBPa were significantly reduced by PC treatment.This result consequently led to suppression of expression of LPL, FABP4,CD36, and SCD2, which are target genes of PPARg and C/EBPa. PC also inhibited protein levels of PPARγ, FABP4, and GLUT4. Accordingly, our results suggest that PC inhibited adipogenesis in 3T3-L1 preadipocytecells through PPARg and CEBPa-related pathways at both transcript and protein levels.Keywords : Phellodendri cortex, 3T3-L1 preadipoctye, Adipogenesis

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Macrocyclic Lactone Derivatives from the Marine-Derived FungusPenicillium sp. Byeng Wha SON*, Zhile FENG, Xavier SIWE-NOUNDOU, Viviane N. NENKEP, Guohua YANG, Alian S. LEUTOU and Keumja YUNDepartment of Chemistry, Pukyong National University, Nam-gu, Busan 608-737, Korea. *Corresponding author: [email protected]

Exploration of structural diversity of marine-derived fungal natural products has been in progress for more than a decade. Early discoveries of the metabolites validated the hypothesis that marine-derived strains offungi could serve as useful sources of unprecedented molecular frameworks. In our studies on the chemistry and biology of fungi, we have investigated secondary metabolite from a marine strain of the fungusPenicillium sp. This strain was isolated from the surface of the drifting cotton clothing collected in Namhae Island, Gyeongnam, Korea and identified based on the morphological evaluation and fatty acid methyl ester analysis (Korean Culture Center of Microorganism, Seoul, Korea, a similarity index of 0.919). Here, we report on the isolation and structural determination of 14-membered resorcyclic acid lactone analogues from a marine isolate of the fungus Penicillium sp. Keywords : Macrocyclic lactone, Fungus, Penicillium sp.

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Effect of High Molecular Weight Poly-γ-glutamic Acid on Moisturizing and Skin Elasticity Chung PARK2, Hyoung Bum KIM2, James TUAN3, Mie IWAMOTO1 IWAMOTO2, Hiroshi UYAMA4, Chul-Joong KIM5, Haryoung POO6 and Moon-Hee SUNG *1,2

1Department of Advanced Fermentation Fusion Science and Technology, KookminUniversity, Seoul 136-702, Korea. 2BioLeaders Corporation, Daejeon 305-500, Korea. 3CPKelco, Atlanta, GA 30339, USA. 4Department of Applied Chemistry, Osaka University, Suita 565-0871, Japan. 5Department of Veterinary Medicine, Chungnam National University, Daejeon. 6Pandemic Influenza Preparedness Research Center, KRIBB, Daejeon 305-333, Korea. *Corresponding author: [email protected]

Ultra high molecular weight poly-γ-glutamic acid (γ-PGA) is a safe and edible biomaterial that is naturally synthesized by Bacillus subtilis (chungkookjang). Development of γ-PGA has pursued in terms of cosmetics, skin care, bone care, nanoparticle for drug delivery system, hydrogel and so on. From the discovery of B. subtilis (chungkookjang) that produces high molecular weight of γ-PGA, specific applications have continued to be developed during the last 10 years. At present, we succeeded in producing γ-PGA with high molecular weight industrially and developing new functional applications in foods and cosmetics. Previous studies showed that γ-PGA could successfully replace hyaluronic aicd and collagen in skin care products. Considerable attention has been focused on the development of anti-aging cosmetics. Very recently we verified the effect of high molecular weight of γ-PGA on moisturizing and skin elasticity. The highlights of these studies and their cosmetics application will be shown and discussed. This work was supported by a Seoul R&BD program (10580)’ in Korea program grant from the Seoul Development Institute. Keywords : poly-γ-glutamic acid, moisturizing and skin elasticity

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Cytotoxic 6-Alkysalicyclic Acids, Salaceyins Inhibits ATPase Activity of Heat Shock Protein 90 Cheng-Zhu WU1,2, An Na MOON3, Oksik CHOI1, Sun-Young KANG1, Jung Joon LEE1, Dongho LEE4, Bang Yeon HWANG5, Young Ho KIM2, Hong-Sub LEE3 and Young-Soo HONG*1

1KRIBB. 2College of Pharmacy ChungNam National University. 3Ildong Pharmaceutical Co. 4Korea University. 5Chungbuk University. *Corresponding author: [email protected]

The molecular chaperone heat shock protein 90 (Hsp90) is responsible for maintaining the correct folding and stability many signaling proteins and is a promising target in cancer and several other diseases, such as neurodegenerative disease, nerve injuries, inflammation, and infection. Inan effort to identify new Hsp90 inhibitors, we attempted to use the ATPaseinhibition assay using Streptomyces culture extracts. Upon screening our extract library, we identified the phenolic lipids, salaceyins A and B as a new Hsp90 inhibitor. Salaceyins A and B were exhibited weak activityof ATPase inhibition with IC50 values of 69.9 and 66.1 μM, respectively.We examined the effects of salaceyin on ATP binding to Hsp90 in vitro by use of ATP-sepahraose. In addition, we will show that salaceyins A and B induced the degradation of Hsp90 client proteins in cancer cells. [This work was supported in part by the 21C Frontier Microbial Genomics and Application Center] Keywords : Hsp90, Streptomyces, inhibitors

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Antioxidant and Antibacterial Activities of Methanol Extracts from Commercial Herbs In-Gyeong CHAE1, Hyun-Jeong KIM2, Mi-Hee YU1, Hyuk Il KIM1 and In-Seon LEE*1

1Department of Food Science and Technology, Keimyung University, Daegu, Korea. 2The Center for Traditional Microorganism Resources, Keimyung University, Daegu, Korea. *Corresponding author: [email protected]

To investigate the antioxidant and antibacterial activities of methanol extracts from 6 herbs which are Chrysanthemum zawadskii Herbich var. latilobum kitam (Siberian chrysanthemum), Lavandula spica L. lavinder (Lavender), Rosmainus offcinals L. (Rosemary) and Cymbopogon citrates (Lemongrass), Saussureae radix (Elecampane), Calendular dffcinalis L (Calendular). Antioxidative effects of herbal extracts were measured by polyphenol, flavonoid contents, DPPH and ABTS radical scavenging activity. We also evaluated the antibacterial activity against Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli. These siberian chrysanthemum, lavender, rosemary and lemongrass extracts showed high polyphenol and flavonoid contents as well as strong DPPH and ABTS radical scavenging activity. In particular, rosemary extract contained highest polyphenol levels as 126.69±2.62 ㎍/mg compared to other herbs.As the DPPH radical scavenging activities, IC50 values of Rosemary were6.23±0.58 ㎍/mL. The rosemary extracts also showed higher antibacterialeffects against S. aureus and E. coli. These results indicate that rosemary could be used as a natural antioxidant and antibacterial agents. Keywords : Antioxdant, Antibacterial, Herbs

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Isolation of a Potent Complement-Activating Polysaccharide from Fruiting Bodies of the Indigenous Korean Mushroom JFMF Sang-Hong YOON*1, Jung-Keun PARK1, Dong-Gwan KIM2, Chang-Muk LEE1, Han-cheol KANG1 and Bon-Sung KOO1

1Functional Material Development Division, National Academy of Agricultural Sciences, Rural Development Administration, Suwon 441-707, Republic of Korea.2Research Center for Drugs, Konkuk University, Seoul 143-701, Korea. *Corresponding author: [email protected]

The anti-complementary substance is able to prevent hemolytic activityby the activation or inhibition of the complement system. Some anti-complementary polysaccharides have been isolated from bacteria, fungi, and plants. In order to develope new mushrooms available as functional food with excellent immunomodulating activity, we investigatedthe anti-complementary activities of water soluble polysaccharides extracted from fruiting bodies of 5 mushrooms which have known as traditional medicinal mushrooms for a long time. It was observed that the crude polysaccharides of JFMF mushroom had a potent anti- complementary activity. Crude polysaccharide of JFMF was isolated into a neutral polysaccharide (JFMF-NP) and three acidic polysaccharides (JFMF-AP1, AP2, AP3) by DEAE-sepharose column chromatography. JFMF-AP2, one of these, showed the very excellent anti-complementary activity of ITCH (inhibition of total complement hemolysis) value 70% on 5 ug/ml concentrations. Its molecular weight was estimated less about5,000 by Toyopearl HW-65F gel filtration chromatography using pullulanmolecular size standard kits. Also, Detection of a cleavage of C3 by immunoelectrophoresis indicated that JFMF-NP and JFMF-AP2 activatesthe complement via both the alternative and classical pathways. Keywords : anti-complmentary activity, polysaccharide, mushroom

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Xanthorrhizol Inhibits and Eradicates Methicilin-Resistant Coagulase-Negative Staphylococcus spp. (MRCNS) Biofilms Yaya RUKAYADI2, Sunghwa HAN3, Dongeun YONG4 and Jae Kwan HWANG*1

1Dept. of Biotechnology, Yonsei University, Seoul 120-749, Korea. 2Yonsei University. 3Dept. of Biomaterials Science and Engineering, Yonsei University, Seoul 120-749, Korea. 4Dept. of Laboratory Medicine & Research Institute of Bacterial Resistance, Yonsei University. Seoul 120-749, Korea. *Corresponding author: [email protected]

MRCNS infections have emerged as a worldwide problem and clinical strains of MRCNS exhibit reduced susceptibility to antimicrobial agents due to their ability to establish biofilm. The aim of this study was to investigate the inhibition and eradication activity of xanthorrhizol against28 isolates of clinical MRCNS. The clinical MRCNS biofilms were formedin the wells of 96-well microtiter plates, in TSB medium. A 0.4% crystal violet was used to stain the biofilms. MICs of xanthorrhizol against 28 clinical MRCNS isolates range 2 - 16 µg/ml and sessile minimum inhibitionconcentration (SMICs) of xanthorrhizol against 28 isolates of clinical MRCNS was 8-fold - to 128-fold (8 - 128 µg/ml) greater than the concentration required to inhibit planktonic growth (MIC). Xanthorrhizolwas able to eradicate the MRCNS bioflms with biofilm eradication concentration (BEC) range 16 - 256 µg/ml. These results suggest that xanthorrhizol has potential to inhibit the growth and eradicate the biofilmformed by clinical MRCNS isolates.Keywords : Biofilm, Methicilin resistant Staphylococcus, Xanthorrhizol

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Isolation and Characterization of six Different Fusaricidins Produced by Paenibacillus Polymyxa E681 and their Possible Role as Elicitors of Induced Systemic Resistance on Plants Seong-Bin KIM, Soo-Young PARK, Soo-Keun CHOI, Choong-Min RYU and Seung-Hwan PARK*Industrial Biotechnology & Bioenergy Research Center, KRIBB, Daejeon, Rep. of Korea. *Corresponding author: [email protected]

Paenibacillus polymyxa E681 produces a lipopeptide antibiotic, fusaricidin, having excellent antimicrobial activity against plant pathogenicfungi and gram-positive bacteria. Fusaricidin is composed of six amino acids residues and contain an unusual fatty acid side chain, 15-guanidino-3-hydroxypentadecanoic acid. In this study, the fusaricidin have been purified from the culture broth of strain E681 by butanol extraction, silica gel and sephadex LH-20 column chromatography. Finally, six fractions containing different fusaricidins with molecular masses of 883, 897(2), 911(2) and 925 Dalton, respectively, were obtained. We analyzed the amino acid composition of each fraction and found that those are fusaricidin A, fusaricidin B, LI-F5a, LI-F5b, LI-F8a and LI-F8b. Besides its known antifungal and antibacterial activities, the fusaricidins were exploited to assess response to plants by examining elicitation of induced systemic resistance (ISR) against a bacterial pathogen in pepper.The purified fusaricidins were drench-applied in the pepper seedlings andtested on ISR against Xanthomoans axonopodis pv. vesicatoria seven days after leaf-infiltration. Even 11 nM fusaricidin application significantly decreased symptom development compared to water control. Keywords : Fusaricidins, lipopeptide, Induced systemic resistance

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Comparison of Functional Effect of Chunggukjang Prepared with Various Soybean Cultivars Hyeong Eun KIM, Min Hwa KIM and Yong Suk KIM*Department of Food science & Technology, Chonbuk National University, Jeonju,Jeonbuk 561-756, Republic of Korea. *Corresponding author: [email protected]

To improve the quality of chunggukjang with better functional effect, such as antioxidant activity, fibrinogen activity, and inhibitory activity against angiotensin converting enzyme, thirty type of chunggukjang wereprepared with different soybean cultivars; large sized soybeans (20 cultivars), small sized soybeans (3 cultivars), and colored soybeans (7 cultivars). Antioxidant activity were determined by DPPH radical scavenging activity, polyphenol contents, and SOD-like activity and thosewere ranged 83.59 (Heukmi) ~ 91.18% (Daeyang), 30.83 (Heukmi) ~ 80.38mg% (Wonkwang), 10.48 (Heukmi) ~ 47.12% (Saeol), respectively. Whenplasmin 1 unit was 100%, fibrinogen activity of chunggukjang prepared with Jinpum#2 and Cheongja#2 were higher 223.89% and 151.29%, respectively, than those of other soybean cultivars. Inhibitory activity against angiotensin converting enzyme showed significant difference among chunggukjang prepared with various soybean cultivars. Those of chunggukjang prepared with Cheongja#2, Sohwang, and Daol were 49.28%, 41.45%, and 38.85%, respectively. From the results, chunggukjang prepared with Wonkwang and Cheongja#2 showed higher functional characteristics than those of other chunggukjang.

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Identification and Production of Antimicrobial Compound fromStreptomyces sp. CS392 Jin Cheol YOO, Seung Sik CHO and Poonam MANDER*Department of Pharmacy, Chosun University, Kwangju 501-759, Republic of Korea,. *Corresponding author: [email protected]

Actinomycetes strain CS392 was found to exhibit broad spectrum of antimicrobial activity against various pathogenic microorganisms. The 16S rRNA sequence revealed that the strain is closely related to Streptomyceslanatus with 98.22% sequence homology. Based on genetic, morphologicaland biochemical characteristic, the strain CS392 was confirmed as belonging to the genus Streptomyces and we named as Streptomyces sp.CS392. The production of antibiotic from the strain was found to be mostfavorable when cultured for 2 days at 28°C and 180 rpm in a shaking incubator using glucose and peptone as respective carbon and nitrogen sources. The antibiotic was extracted using ethylacetae (1:1 v/v) and partially purified using both column and thin layer chromatographic techniques. Two antibiotic compounds, based on its different solubility on ethyl acetate and methanol, were identified. Antimicrobial activity of the compound against different pathogenic strains was assessed using the minimum inhibitory concentrations (MICs). It was found to be active against eight different strains including MRSA 693E , MRSA (2-32), MRSA (4-5), MRSA (4-21), Enterococcus faecalis (ATCC 29212), Bacillus subtilis (ATCC 1928), Staphyloccus aureus (KCTC 1928) and Mycobacterium smegmatis (ATCC 9341). Keywords : Actinomycetes, Streptomyces sp. CS392, antibiotic

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PTP1B Inhibitory Secondary Metabolites from Antarctic Lichens Myeong-Sook KANG1, Changon SEO1, Joung Han YIM2 and Hyuncheol OH*1

1Dept. of Bio-Food Materials, Silla University, Busan 617-736, Korea. 2Korea Polar Research Institute, KORDI, Incheon 406-840, Korea. *Corresponding author: [email protected]

In the course of screening for PTP1B inhibitory natural products, the MeOH extract of the dried sample of Antarctic lichen Cladonia borealis was subjected to C18 functionalized silica gel flash column chromatography, eluting with a stepwise gradient consisting of MeOH in H2O. The fraction eluted at 100% MeOH was then subjected to semi- preparative reversed-phase HPLC to yield methyl 4-O-demethylbarbatate (1). The MeOH extracts of Antarctic lichen Psudephebe pubescens and Stereocaulon alpinum were also investigated to yield 5,7-dihydroxy- 4-methylphthalide (2) and bourgeanic acid (3), respectively. The structuresof the isolated compounds were identified by analysis of NMR and MS data, along with comparison with those in the literatures. These compounds inhibited PTP1B activity with 50% inhibitory concentration values of 9.72 μM, 100.37 μM, and 5.62 μM, respectively. In addition, the characteristicsof the inhibition of PTP1B by bourgeanic acid (3) were analyzed. Details of the isolation, structure determination, and their PTP1B inhibitory activities will be presented. Keywords : Antarctic lichens (Cladonia borealis, Psudephebe pubescens,

and Stereocaulon alpinum), Depside and Benzofuranoid, PTP1B

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Secondary Metabolites from Antarctic Lichen Lecidella Carpathicaand their PTP1B Inhibitory Effect Changon SEO1, Chi-Wook LEE1, Joung Han YIM2 and Hyuncheol OH*1

1Dept. of Bio-Food Materials, Silla University, Busan 617-736, Korea. 2Korea Polar Research Institute, KORDI, Incheon 406-840, Korea. *Corresponding author: [email protected]

In the course of screening for PTP1B inhibitory natural product, the MeOH extract of the dried sample of Lecidella carpathica was found to exhibit significant inhibitory effect.The MeOH extract was subjected to C18 functionalized silica gel flash column chromatography, eluting with a stepwise gradient consisting of MeOH in H2O. The fraction eluted at 100% MeOH was then subjected to semi-preparative reversed-phase HPLC to yield compounds 1-3. The fraction eluted at 50% MeOH in CH2Cl2 was then subjected to semi-preparative reversed-phase HPLC to yield compound 4 and 5. Compounds 1-5 were identified as atraric acid (1), methyl oresllinate (2), brialmontin 1 (3), atranorin (4), and 6,22-hopanediol (5) by analysis of NMR and MS data, along with comparison with those in the literatures. These compounds inhibited PTP1B activity with 50% inhibitory concentration values of 51.5 μM, 59.5 μM, 14 μM, 10 μM, and 3.7 μM, respectively. Furthermore, the kinetic analysis of PTP1B inhibition by brialmontin 1 (3) and 6,22-hopanediol (5) suggested that these compounds inhibited PTP1B activity in a competitive manner. Details of the isolation, structure determination, and their PTP1B inhibitory activities will be presented. Keywords : Antarctic lichen (Lecidella carpathica), Depside and Terpenoid,

PTP1B

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Optimization for Lacticin BK6 Production by Bacillus Vallismortis BK6 Isolated from Jeot-gal. Ik Soo JANG2, Dong-hwi KIM1, Bong-kun JHON3, Chang-young SONG2

and Moon-soo HEO*1

1Dept. Marine Biomedical Sciences, jeju-University, Jeju 690-120, Korea. 2Dept. Marine life Science, jeju-University, Jeju 690-120, Korea. 3Dept. Marine life Sciences ,jeju-University, Jejul 690-120, Korea. *Corresponding author: [email protected]

Jeot-kal, is a salted termented food in Korean cuisine. It is made with various seafood, such as shrimp, oysters, shellfish, fish, fish egg, and fishintestines. This productis usually fermented using certain types of microorganism sand enzymes. This is not only as a source of protein supply, due to its peculiar flavor, and relative abundance of free amino acid and taste related components, but also a subsidiary material of Kimchior spices.Six bacterial strains were isolated from several Jeot-kal using MRS agar medium. A bacterium which has high activities such as protease and Pathogens bacterial species, was isolated from Myungran Jeot-kal, which was recorded as Bacillus valismortis BK6. It was identified as Bacillus Vallismortis BK6 as based on the biochemical properties and 16S rRNA gene sequence. The bacterium was Gram-positive, catalase- negative, oxidase-positive, hydrosis of esculin ferric citrate, was rod- shaped, and had dimensions of 0.6∼0.7㎛ and 1.5∼2.0㎛. The strains showed characteristics Secrete substance of gooey. When the material was rubbed it was slippery.Nutritional and cultural conditions for the productionof antibacterial substance by this organism under shake-flask conditions have optimized. Keywords : Bacillus valismortis BK6, Cultural Characteristics, Jeot-kal

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A Method for Screenig of Polyene Producing Actinomycetes Jin-Yong KIM, Hyung-Jin KWON and Joo-Won SUH*Department of Biological Science, Division of Bioscience and Bioinformatics, Myongji University, Yongin 449-728, Korea. *Corresponding author: [email protected]

The amphotericin B, a typical polyene type antifungal compound, has widely used as a clinical fungicide agent for treatment of systemic infection. However, it has been reported that treatment of amphotericin B was not successful in many cases. That is not because of resistance by infected pathogen but hypofunction by host. Therefor, a few amphotericin B resistance pathogen was reported. So, polyene type antifungal compounds such a amphotericin B are more clinically practical than other type of antifungal compounds like azole and 5-fluorocytosine. The amphotericin B can incorporate with sterol and ergostreol in phospholipid membrane of fungal and form ionic membrane channels, which cause the loss of barrier function in fungal membrane, raising fungal cell death. Based on this mechanism, we hypothesized that the amphotericin B activity in broth culture of Candida albicans ATCC10231,a typical human pathogen, should be reduced with optimal concentrationof ergosterol. So, we determined 50% inhibitory concentration(IC50) of the amphotericin B against Candida albicans ATCC10231 as 0.05㎍/㎖

and optimized the concentration of ergosterol in IC50 of the amphotericinB as 5㎎/㎖. By using this method, we successfully screened several polyene producing Actinomycetes. Keywords : screening for antifungal compounds, polyene producing

Actinomycetes, amphotericin B

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Biosynthesis of Polymyxin using Bacillus Subtilis as a Surrogate Host Soo-Young PARK , Soo-Keun CHOI, Seong-Bin KIM, Jihyun F. KIM and Seung-Hwan PARK*Industrial Biotechnology & Bioenergy Research Center, KRIBB, Daejeon, Rep. of Korea. *Corresponding author: [email protected]

Polymyxin, a long-known peptide antibiotic, has recently been reintroduced in clinical practice because it is the only available antibioticfor the treatment of multidrug-resistant Gram (-) pathogenic bacteria. We identified a gene cluster encoding polymyxin synthetase by whole genomesequencing of Paenibacillus polymyxa E681. The gene cluster spanning the 40kb region consists of 5 open reading frames, pmxABCD, and pmxE. The pmxC and pmxD genes are similar to genes encoding transport proteins, and pmxAB and E encode polymyxin synthetase, nonribosomal peptide synthetic mode. Also, we found that the modular structures of polymyxin synthetases identified from three different P. polymyxa strainsproducing polymyxin A, B, and E, respectively, are very similar to one another. The pmx gene cluster was introduced into B. subtilis chromosome. And then, to produce polymyxin successfully without exogenous L-2,4-diaminobutyric acid (DAB) which is consist of major amino acid of polymyxin absent in B. subtilis, ectB encoding DAB synthetase was introduced. As expected, the resulting strain could produce polymyxin successfully without exogenous DAB. We have also found that AbrB, a transition state regulator of B. subtilis negatively affects the expressionof the polymyxin synthetase gene. Keywords : Paenibacillus polymyxa, Polymyxin, Bacillus subtilis

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Screening of Enhanced Bioactivity in Biotransformed Morus albaL by Commercial Enzyme Sang Hun KIM, Sun Nyoung YU, Kwang Youn KIM, Dong Min KANG, Young Rang JIN, Hyun Joo JEON and Soon Cheol AHN*Department of Microbiology and Immunology, Pusan National University Schoolof Medicine, Beomeo-ri, Mulgum-eop, Yangsan 626-770, Gyeoungnam, Korea. *Corresponding author: [email protected]

Morus alba L, contains many bioactive components (flavones, triterpenes) and possess several biological effects (antioxidative, antiinflammation), has been used as folk medicine and traditional beverage in Korea. We tried to enhanced biocompound of muberry leave through biotransformation.In this study, muberry leaves were transformed to partialdegradation by commercial enzymes such as protease, lipase and carbohydratases. We confirmed its degradation and newly synthesis compounds by TLC and investigated the bioactivities such as the antioxidant effect, the cytotoxicity effect, the mushroom tyrosinase inhibition activity, the pancreatic lipase inhibition activity, the nitric oxide effect.In the present study, the biotransformed extract of meberry leaves were found to have high radical scavenging activity, cytotoxicity, tyrosinase activity and lipase activity in transforment by AMG and Viscozyme, but nitric oxide activity were not significant found the high activity from total extract. Taken together, we confirmed its modification by biotransformation and confirmed increased bioactivity though various assay. So, we suggest that it is possible to use as functional foods or dietary supplements. Keywords : Moberry alba L, Biotansformation, Bioactivity

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Characterization of Anti-angiogenic Activities of Carnosol, an Active Principle of Rosemarinus Officinalis Seung Wook KIM1,2, Ki Hyun KIM1, Hyung Keun LEE1, Seung Hyeon JUNG2 and Ho Jeong KWON*1

1Department of Biotechnology, Translational Research Center for Protein Function Control, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749, Korea. 2Ottogi Central Research Center, Ottogi Corporation, Anyang, Kyeonggi-Do, Korea. *Corresponding author: [email protected]

Spice and herbs are used in many foods and functional foods for their biological effects. Accordingly, we have investigated the inhibition activityof spice and herb extracts on aminopeptidase N (APN) which plays a crucial role in angiogenesis. Rosemary (Rosemarinus Officinalis, Labiatae)extracts have showed a relatively high inhibitory activity toward APN. Rosemary extract has been known to contain rich antioxidants. Most of these antioxidant activities are contributed by carnosol and carnosic acids.Carnosol is a naturally occurring phenolic diterpene and revealed that has various medicinal effects including anti-acncer and anti-inflammatory activities in various disease models. Based on this idea, we focused on anti-angiogenic activity of carnosol. A cell proliferation assay was conducted on various cell lines. Carnosol showed relatively stronger inhibition for growth of human umbilical vein endothelial cells (HUVECs)than that of cancer cell lines. Carnosol potently inhibited VEGF-induced angiogenesis of endothelial cells in vitro and in vivo angiogenesis of chorioallantoic membranes from growing chick embryos. Taken together, carnosol can be developed as a potent anti-angiogenic agent. Mode of actions of carnosol on anti-angiogenic activity are under investigation.Keywords : angiogenesis, aminopeptidase N (APN), carnosol

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The Protective Effects of Exo-biopolymer from Submerged Cultureof Lentinus edodes with Rice Bran on Acute Ethanol-induced and CCL4-induced Hepatotoxicity of Rats Seonggil HONG, Joong-Hark KIM, Hwa-Young KIM, Dong Ju KIM and Sung-Joo HWANG*Erom R&D center, Erom Co. Ltd, Chungcheon, Korea. *Corresponding author: [email protected]

Previous studies have shown that exo-biopolymer from submerged culture of Lentinus edodes with rice bran have various kinds of health beneficial effects such as anti-cancer activity, immune enhancement and antioxidation activity. And also beta-glucan, one of the polysaccharide contained in rice bran and Lentinus edodes was suggested to prevent andenhance obesity and obesity related diabetes through antioxidative activity and improvement of lipid metabolism in recent researches. In the preliminary study, we were development of the culture condition for exo-biopolymer production through secondary fermentation with Lactobacillus. In this study, we were evaluate the protective effects of existing Exo-biopolimer(RBF) and improving Exo-biopolimer by secondary fermentation with Lactobacillus(RBFLB) on acute hepatotoxicity inducedby ethanol and carbon-tetracycline in rats. In ethanol-induce model, rats were administered each substance during 2 weeks and CCL4-induced model were 4 weeks. We measured the concentration of glutamate-oxalatetransaminase and glutamate-pyruvate transaminase in serum to observe the damage to liver. And serum and hepatic antioxidation relating factorswere measured to evaluate reducing effects to hepatotoxicity of substance. Keywords : bio bran, hepatotoxicity, lentinus edodes

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Glucopyanosylspinasterol from the Leaves of Stewartia Koreanaand its Inhibitory Effects on UVB-induced Collagen Reduction and Matrix Metalloproteinase-1 Gene Expression in Human Dermal Fibroblast Sang Min LEE, Tae Hoon LEE, Myun-Ho BANG, Nam-In BAEK and Jiyoung KIM*College of Life Science and Graduate School of Biotechnology, Kyung Hee University, Yongin 446-701, Korea. *Corresponding author: [email protected]

We identified a number of bioactive compounds from the leaves of Stewartia koreana and examined the effect of one of active compounds, 3-O-β-D-glucopyanosylspinasterol (spinasterol-Glc) on the levels of collagen and MMP-1 in UVB-irradiated fibroblast cells. Methanol fractionof the leaves of Stewartia koreana was partitioned with EtOAc, n-BuOH,and H2O, successively. The repeated SiO2 and ODS column chromatography for the EtOAc fraction led to isolation of one of the active compounds. Treatment of human dermal fibroblast cultures with spinasterol-Glc led to restore UVB-mediated reduction of collagen in the fibroblast cells in dose-dependent manner. The compound inhibited expression of MMP-1 in UVB-treated fibroblast cells at the mRNA and protein levels, while it enhanced expression of TIMP-1 and TIMP-2, inhibitors of metallopro-teinases. It was further demonstrated that spinasterol-Glc inhibited IkB-αphospholylation and IkB-α degradation, resulting in the inhibition of UVB-induced translocation into nucleus and DNA binding to its target sites of NF-kB. The results demonstrate that spinasterol-Glc inhibits collagen degradation via inhibition of MMP-1 gene expression in UVB-induced fibroblast cells. Keywords : Glucopyanosylspinasterol, collagen reduction, matrix

metalloproteinase-1

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Effects of Deinoxanthin in PC-3 Human Prostate Cancer Cells Hyun-Pa SONG, Sang-Yong LIM, Min-Ho JOE, Eun-Hye SHIN and Dong-Ho KIM*Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute. *Corresponding author: [email protected]

Effects of deinoxanthin, major pigment of D. radiodurans R1, on PC-3 human prostate cancer cells were investigated. The crude extract of carotenoids from D. radiodurans R1 was analyzed using high performanceliquid chromatography (HPLC) and exhibited more than 5 peaks with one major pigment. Deinoxanthin, major pigment, was purified with a purityof 95% using semipreparative HPLC. Confirmation of the identity of deinoxanthin was provided by HPLC-atmospheric pressure chemical ionization-mass spectrometry (APCI-MS) analysis in the positive ionization mode. Treatment of PC-3 cells with deinoxanthin resulted in a dose-dependent inhibition of cell viability and induction of apoptosis which was associated with the generation of reactive oxygen species (ROS). Exposure of PC-3 cells to deinoxanthin also associated with increased expression of Bax, down-regulation of Bcl-2, and significant release of cytochrome c into the cytosol. In addition, deinoxanthin treatment led to the activation of caspase-9 and -3, accompanied by the cleavage of poly (ADP-ribose) polymerase (PARP). These results suggestthat treatment of human prostate cancer cells with deinoxanthin induced apoptosis in a dose-dependent manner and that this effect was mediated by generation of ROS. Keywords : Deinococcus radiodurans R1, Deinoxanthin, Prostate cancer

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A Study of Functional Materials Using the Extract from Hen’Eggshell Membrane Bit-Na KIM1, Yang-Hoon KIM2 and Jiho MIN*1

1Graduate School of Semiconductor and Chemical Engineering, Chonbuk National University. 2School of Life Science, Chungbuk National University. *Corresponding author: [email protected]

Eggshell membrane (EM) is a novel, robust, microporous, cost effective, easily available organic materials. In this study, EM was chemically or physically treated to obtain functional materials containing several organicacids and proteins as a medium supplements to stimulate bacterial cell growth. Many solvents were tested for solubilization of Hen’s EM, and buthanol, sodium hydroxide, potassium hydroxide, respectively dissolved EM as a solution type, but it was not showed to increase final cell densityand growth rate of Escherichia coli grown under each supplemented condition using above mentioned solutions. Therefore, functional materials extracted from EM were obtained using physical extraction methods, i.e.direct autoclave and sonication of EM, and then applied to stimulate E. coli growth. In addition, the comparison to adding of glucose and EM extract, the results indicated that EM was not a carbon source, but physiological activating nutrient on the growth of microorganism. In the E. coli, the growth which was faster about 2.7 times at the lag phase. Overall result strengthens our view of using physically extracted EM as physiological activating materials for bacterial cell growth stimulation and can be used in various industrially microbial applications. Keywords : Eggshell membrane

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Potent Anticariogenic Activity of Aceriphyllum Rossii and its cOmponents, Aceriphyllic Acid A and 3-oxoolean-12-en-27-oic acid Yun-ju KWON, Chang-Ji ZHENG, Hyun-Woo OH and Won-Gon KIM*Korea Research Institute of Bioscience & Biotechnology, Yusong, Daejeon 305-806, Korea. *Corresponding author: [email protected]

Aceriphyllum rossii Engler (Saxifragaceae) have been used as a nutritious food in Korea. We found that the methanol extract of A. rossii root and its components, aceriphyllic acid A and 3-oxoolean-12-en-27-oic acid, potently inhibited the growth of the key cariogenic bacteria, Streptococcus mutans, with MIC of 2 - 4 µg/mL. They also showed antibacterial activity against other cariogenic bacteria such as S. oralis, S. sobrinus, and S. salivarius with the similar potency. In the time-kill study, aceriphyllic acid A reduced the viable counts of S. mutans by 90% in 1 min at 8 mg/mL, indicating that aceriphyllic acid A had the fast bacteriostatic activity. Severe damages of the cell surface of S. mutans by aceriphyllic acid A were observed by transmission electron microscopy,suggesting with its fast antibacterial activity that its mechanism of actionmight be membrane disruption. These results suggest that the methanol extract of A. rossii root and its components, aceriphyllic acid A and 3-oxoolean-12-en-27-oic acid, could have the great potential as natural agents for preventing dental caries. Keywords : Streptococcus mutans, Aceriphyllum rossii, anticariogenic

activity

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Methyl-Branched Fatty Acids, Inhibitors of Enoyl-ACP Reductasewith Antibacterial Activity from Streptomyces sp. A251 NYUNG KIM, Chang-Ji ZHENG, Mi-Jin SOHN, Seung-Wook CHI and Won-Gon KIM*Korea Research institute of Bioscience & Biotechnology, Yusong, Daejeon 305-806, Korea. *Corresponding author: [email protected]

Bacterial enoyl-ACP reductase (FabI) has been demonstrated to be a novel antibacterial target. In the course of our screening for FabI inhibitorswe isolated two methyl-branched fatty acids from Streptomyces sp. A251.They were identified as 14-methyl-9(Z)-pentadecenoic acid and 15-methyl- 9(Z)-hexadecenoic acid by MS and NMR spectral data. These compoundsinhibited Staphylococcus aureus FabI with IC50 of 16.0 and 16.3 mM, respectively, while didn’t affect FabK, an enoyl-ACP reductase of Streptococcus pneumonia, at 100 mM. Consistent with their selective inhibition for FabI, they blocked intracellular fatty acid synthesis as wellas the growth of S. aureus, while didn’t inhibit the growth of S. pneumonia. Additionally, these compounds showed reduced antibacterial activity against fabI-overexpressing S. aureus compared to the wild-type strain. These results demonstrate that the methyl-branched fatty acids showed antibacterial activity by inhibiting FabI in vivo.. This work was supported by the 21C Frontier Microbial Genomics and Application Center Program.Keywords : methyl-branched fatty acid, enoyl-ACP reductase, antibacterial

activity

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Isolation and Identification of Pentagalloylglucose with Broad-Spectrum Antibacterial Activity from Rhus Trichocarpa Miquel NYUNG KIM, Jun-Young CHO, Mi-Jin SOHN, Joong-ku LEE and Won-Gon KIM*Korea Research Institute of Bioscience & Biotechnology, Yusong, Daejeon 305-806, Korea. *Corresponding author: [email protected]

Rhus trichocarpa Miquel has been utilized both as a food and for medicinal purposes. In this study, we determined that the methanol extracts of the stem and leaf portions of R. trichocarpa inhibited the growth of both Gram-negative and Gram-positive bacteria. The active constituent wasisolated, and identified via MS and NMR as 1, 2, 3, 4, 6-penta-O- galloyl-b-D-glucose. The compound also evidenced a broad spectrum of antibacterial activity against both Gram-negative and Gram-positive bacteria including MRSA, QRSA, B. subtilis, S. mutans, E. coli, P. aeruginosa and S. typhimurium with MIC values of 16 - 32 µg/ml, whereasgallate failed to inhibit the growth of Gram-negative bacteria, even at a concentration of 128 µg/ml. The antibacterial activity of penta-O- galloylglucose was restored by the addition of Fe2+, whereas gallate wasn’t, thereby indicating that its antibacterial activity could be attributableto the chelation of iron. The results of time-kill study against S. aureus and E. coli revealed that penta-O-galloylglucose exhibited bacteriostatic activity. These findings indicate that the extracts of R. trichocarpa as wellas its active component, penta-O-galloylglucose, may have profound potential for the control of both Gram-positive and negative pathogens. Keywords : Rhus trichocarpa, broad spectrum antibacterial activity,

penta-O-galloylglucose

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Production of Plant-Specific Naringenin by Escherichia coli Containing an Artificial Synthetic Genes Oksik CHOI1,2, Cheng-Zhu WU1, Sun Young KANG1, Jung Joon LEE1, Tai-Boong UHM2 and Young-Soo HONG*1

1KRIBB. 2Chonbuk National University. *Corresponding author: [email protected]

Polyphenols produced in a wide variety of plants have the potential to be valuable fine chemicals for the treatment of an assortment of humandiseases. One of the major constituents within this chemical class is flavonoids, among which flavanones as the precursor to all flavonoid structures, are the most prevalent. Flavanone (naringenin), the central precursor of flavonoids, are synthesized exclusively in plants from tyrosinevia the sequential reaction of tyrosine ammonia-lyase (TAL), 4-coumarate: coenzyme A ligase (4CL) and chalcone synthase (CHS). For the purpose of construction of artificial metabolic pathway which contained four genes of heterologous origins, tyrosine ammonia lyase (TAL) from Saccharothrix espanaensis, cinnamate/coumarate:CoA ligase (CCL) from Streptomyces coelicolor, chalcone synthase(CHS) from Arabidopsis thaliana, and chalcone isomerase(CHI) from Medicago sativa were constructed. The artificial synthetic pathway in E. coli shows an efficient production of naringenin. This artificial biosynthesis is a new tool for the production in bacteria of rare and expensive plant and animal natural products. [Thiswork was supported in part by the 21C Frontier Microbial Genomics andApplication Center] Keywords : polyketide, synthetic biology, biosynthesis

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A Curcumin Analog HBC Inhibits Angiogenesis by Downregulating Hypoxia-Inducible Factor Hye Jin JUNG, Jong Hyeon KIM, Joong Sup SHIM and Ho Jeong KWON*Dept. of Biotechnology, Yonsei University, Seoul 120-749, Korea. *Corresponding author: [email protected]

Recent reports have shown that Ca2+/calmodulin (Ca2+/CaM) signalingplays a crucial role in angiogenesis. We previously developed a new Ca2+/CaM antagonist, HBC (4-{3,5-Bis-[2-(4-hydroxy-3-methoxy- phenyl)-ethyl]-4,5-dihydro-pyrazol-1-yl}-benzoic acid), from the curcumin based synthetic chemical library. Here, the anti-angiogenic activity of HBCwas investigated. HBC potently inhibited the proliferation of human umbilical vascular endothelial cells with no cytotoxicity. Furthermore, HBC blocked in vitro angiogenesis such as tube formation and chemoinvasion as well as neovascularization of the chorioallantoic membrane of growing chick embryo in vivo. We also evaluated the effect of HBC on the activity of hypoxia-inducible factor (HIF) which plays a critical role in hypoxia-induced angiogenesis. The compound markedlyinhibited HIF-1a expression at the translational level during hypoxia, thereby reducing HIF-1 transcriptional activity and expression of its major target gene vascular endothelial growth factor. Collectively, HBC is a promising anti-angiogenic agent targeting HIF and can be utilized to explore the biological role of Ca2+/CaM in angiogenesis. Keywords : Ca2+/calmodulin, HBC, angiogenesis

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In vitro and In vivo Antifungal Activity of Bacillus velezensis G341 against Various Phytopathogenic Fungi and Characterizationof Its Antifungal Components Mi-Young YOON1,2, Myung Soo PARK1, Won Ki KIM1, Gyung Ja CHOI1,Kyoung Soo JANG1, Yong Ho CHOI1, Young-Ho KIM3, ByeongJin CHA2

and Jin-Cheol KIM*1

1Chemical Biotechnology Research Center, Korea Research Institute of ChemicalTechnology, Daejeon 305-600, Korea. 2College of Agricultural and Life Sciences,Chungbuk National University, Cheongju, Chungbuk 361-763, Korea. 3Departmentof Agricultural Biotechnology, Seoul National University, Seoul, 151-921, Korea. *Corresponding author: [email protected]

The Bacillus velezensis G341, which was isolated from the four-year-old roots of Korean ginseng with rot symptoms, showed strong antifungal activity against various phytopathogenic fungi. In in vitro assay using Magnaporthe oryzae, its optimal medium was Tryptone Soya Broth (TSB) medium with regard to produce antifungal substance. Its TSB fermentation broth was also highly effective in controlling rice sheath blight. The antifungal activity of the culture filtrate was significantly correlated with the cell growth of strain G341. The active metabolite in the culture filtrate was relatively thermally stable with antifungal activity of the culture filtratebeing retained even after being held at 121oC for 30 min. Meanwhile, the antifungal activity of the culture filtrate against the growth of M. oryzae remained almost unchanged when the culture was exposed to a pH ranging from 6 to 10. Strong in vivo antifungal activity was shown in 1-butanol fraction in in vivo bioassay. The antifungal compounds were identified as bacillomycin L and fengycin. The two compounds also showed specifically potent in vivo antifungal activity against rice sheath blight. This study highlights biocontrol potential of strain G341 producing bacillomycin L and fengycin in management of rice sheath blight. Keywords : Bacillus velezensis G341, bacillomycin L, fengycin

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PTP1B Inhibitory Effect of Botcinin B Isolated from Marine-derived Fungus SF-5275 Min-Young KIM1, Jae Hak SOHN1, Jong Seog AHN2 and Hyuncheol OH*1

1Dept. of Bio-Food Materials, Silla University, Busan 617-736, Korea. 2Korea Research Institute of Bioscience and Biotechnology (KRIBB), 52 Eoun-dong, Yuseong-gu, Daejeon 305-333, Republic of Korea. *Corresponding author: [email protected]

To identify PTP1B inhibitors from the fungal strain SF-5275, the organic extract from the fungal culture was fractionated and purified by utilizing the solvent extraction, reversed-phase flash column (C18) chromatography,and HPLC. The bioassay-guided fractionation and purification afforded two related fungal metabolites 3-O-acetyl botcinic acid methyl ester (1) and botcinin B (2). The structures of these compounds were mainly determined by analysis of NMR and MS data. These compounds inhibited PTP1B activity with the 50% inhibitory concentration values of 210 μM and 53.6 μM, respectively. Furthermore, the kinetic analysis of PTP1B inhibition by compound 2 suggested that the compound inhibited PTP1B activity in a non-competitive manner. Although botcinin B has been recognized as an antifungal agent, its moderate PTP1B inhibitory effect was identified for the first time in this study. Details of the isolation, structure determination, selectivity of other PTP1B enzymes and their PTP1B inhibitory activities will be presented. Keywords : Marine Fungus, Botcinin B, PTP1B

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Triherbal Enriched Diet on Physiology and Immunology of Paralichthys Olivaceus Infected with Streptococcus Iniae Ik Soo JANG2, Ramasamy HARIKRISHNA2, Ju-Sang KIM2, Kyung-Mi KIM1 and Moon-Soo HEO*1

1Dept. Marine Biomedical Sciences, jeju-University, Jeju 690-120, Korea. 2Dept. Marine life Science, jeju-University, Jeju 690-120, Korea. *Corresponding author: [email protected]

This study determines the effect of dietary mixed herbal supplemented feed in Paralichthys olivaceus against Streptococcus iniae , through the changes in hematological, biochemical, and immunological parameters observed for 8 weeks. The WBC level of infected untreated and triberbaldiets groups significantly decreased throughout the experimental trial while it was no change in H group after weeks 6 to 8 compared to 0 week. The haemoglobin and haematocrit levels significantly decreased in all thegroups compared to 0 week, except in group fed with 100 mg kg-1 of mixed herbal supplemented diet on week 6 to 8. The mean corpuscular volume and mean corpuscular haemoglobin values did not change in controland H groups at any sampling compared to 0 week. The mixed herbal diet feeding group did not result in any change in the percentage of lymphocytes after week 6 and 8. The monocytes and eosinophils did not change in C and H groups. The study indicates that administration of mixed herbal supplemented feed was effective in restoring the affected hematological and biochemical parameters and enhanced the nonspecific immune system of P. olivaceus infected with S. iniae. Keywords : immunology, Paralichthys olivaceus, Streptococcus iniae

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Methoxymalonyl-acyl Carrier Protein Biosynthesis Locus, While Dispensable for the Galbonolide B Production, is Essential to the Biosynthesis of Galbonolide A in Streptomyces Galbus Hyung-Jin KWON*1, Suman KARKI1, Hae-Kyung YOO1, So-Yeon KWON1

and Si-Hyung PARK2

1Division of Bioscience and Bioinformatics, Myongji University. 2Major in Medicinal Plant Resources, Division of Bioscience, Mokpo National University. *Corresponding author: [email protected]

Galbonolide A and B are antifungal compounds, which are produced by Streptomyces galbus. A multi-modular polyketide synthase (PKS) was predicted to catalyze their biosynthesis, and a methoxymalonyl-acyl carrierprotein (-ACP) was expected to be involved in the biosynthesis of galbonolide A. Cloning of a methoxymalonyl-ACP biosynthesis locus (galGHIJK) and the flanking regions revealed that the locus was co-localized with β-ketoacyl synthase (KAS)-related genes (orf3, 4, and 5) but separate from any multi-modular PKS gene cluster in S. galbus. A galI-disruption mutant (SK-galI-5) is unable to produce galbonolide A,but can synthesize galbonolide B, indicating that galGHIJK is involved in the biosynthesis of galbonolide A. A disruption mutant of orf4 is severelyimpaired in the production of both galbonolide A and B. These results indicate that galGHIJK and the KAS genes are involved in the biosynthesis of galbonolides, although they are not co-localized with a multi-modular PKS gene cluster. We further propose that a single galbonolide PKS generates two discrete structures, galbonolide A and B, by alternatively incorporating methoxymalonate and methylmalonate, respectively. Keywords : galbonolide, methoxymalonyl-ACP, Streptomyces galbus

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A Functional Study on Lysosome-Related Organelles Extract for Multipurpose Eco-materials Jihee YOON1, Yang-Hoon KIM2 and Jiho MIN*1

1Division of Semiconductor and Chemical Engineering, Chonbuk National University, 664-14 Duckjin-dong, Jeonju 561-756, South Korea. 2School of Life Science, Chungbuk National University 410 Sungbong-Ro, Heungduk-Gu, Cheongju 361-763, South Korea. *Corresponding author: [email protected]

Lysosomes are heterogeneous cell organelles consisting of vacuoles that vary in size, form, and density, and lysosomes release lysosomal enzymesto digest intracellular and extracellular waste material. Lysosomal enzymes degrade bacteria cell walls, demonstrating the antimicrobial activity of lysosomes. Lysosomes are not silent organelles, but dynamic organelles in which lysosomal enzymes are easily integrated or released when exposed to stressful conditions, especially oxidative stress. In addition, it was reported that disrupted lysosomal function over aging may further contribute to pigment accumulation. Therefore, in this study, we isolatedlysosome-related organelles from Saccharomyces cerevisiae, HeLa cell lines and Hen’s egg white, and then used organelles extract for the reduction of melanin causing hypermelanosis. We found that as a periodof 20 days, 90% of melanin was reduced by treatment of organelles extract,as measured by the reduction of melanin pigmentary intensity. Therefore, our results provide insight for the application of in vitro lysosomes as antimicrobial agents and as apoptosis-inducing materials, as well as usefulagents for cosmeceutical skin lightening and treatment of hyperpigmenta-tion disorders. Keywords : Lysosomes, cell organelle, melanin, Antimicrobial,

anticancerous agent

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Inhibition of Atopic Dermatitis-like Skin Lesions by Topical Application of Cedrol Jin Hee KIM*1, Yeo Dae YOON2, Eun Sook LEE1

1Dept. of Herbal Skin Care, Daegu Hanny University, Gyeongsan 712-715, Korea.2Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-333, Korea. *Corresponding author: [email protected]

Atopic dermatitis (AD) is a chronic inflammatory skin disease that commonly begins in childhood. Cedrol, a sesquiterpene isolated from the leaves of Chamaecyparis obtusa, exhibits with inhibition of atopic dermatitis-like skin lesions. In the study, we examined the effect of topicalapplication of cedrol on skin inflammation and AD-like skin lesions in mouse models. Cedrol dose-dependently inhibited phorbol ester-induced increase in ear thickness in BALB/c mice. Histopathological analysis revealed that ear swelling was suppressed by cedrol treatment. Also, cedrol inhibited interleukin-6 (IL-6) and tumor necrosis factor-a (TNF-a) secretion from the phorbol 12-myristate 13-acetate and A23187-induced HMC-1 cells (human mast cell line). Furthermore, cedrol suppressed the mRNA expression of IL-6 and TNF-a in phorbol 12-myristate 13-acetate and A23187-induced HMC-1 cells.Taken together, our results showed that topical application of cedrol exerts beneficial effects in animal model of skin inflammation and AD, suggesting that cedrol might be a promising topical agent for the treatment of inflammatory skin diseases. Keywords : Atopic dermatitis, Cedrol

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Identification and Characterization of Regulatory Network of Tautomycetin from Streptomyces sp. CK4412 Jihye NAH, Si-Sun CHOI and Eung-Soo KIM*Department of Biological Engineering, Inha University, Incheon 402-751, Korea.*Corresponding author: [email protected]

Tautomycetin (TMC), which is produced by Streptomyces sp. CK4412, is a novel activated T cell-specific immunosuppressive compound with an ester bond linkage between a terminal cyclic anhydride moiety and a linear polyketide chain bearing an unusual terminal alkene. Previously,we isolated and characterized the entire TMC biosynthetic gene cluster from Streptomyces sp. CK4412. Within the TMC biosynthetic gene cluster, we found two putative pathway-specific genes, tmcN and tmcT. Both TmcN and tmcT were identified as positive regulators by targeted gene disruption as well as in silico sequence comparison. Here, we isolated, identified, and disrupted a wblA ortholog gene (named wblA-tmc) from Streptomyces sp. CK4412, which was previously reported as a strong global down-regulator of antibiotic biosynthesis in various Streptomycetesspeices. Moreover, we constructed two double knock-out strains such as △tmcT△wblA-tmc as well as △tmcN△wblA-tmc in Streptomyces sp. CK4412 to understand how a global regulatory gene wblA-tmc regulates TMC pathway-specific regulatory tmcN and/or tmcT genes in a TMC regulatory network in Streptomyces sp. CK4412. The detailed results willbe further discussed. Keywords : Streptomyces, Immunosuppressant, Tautomycetin

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Suppression of Rice Blast and Pepper Phytophthora Blight by Solid-state Fermentation Cultures of Bacillus velezensis G341Jin-Cheol KIM*, Won-Ki KIM, Myung Soo PARK, Gyung Ja CHOI, Yong Ho CHOI and Kyoung Soo JANGChemical Biotechnology Research Center, Korea Research Institute of Chemical Technology, Daejeon 305-600, Korea. *Corresponding author: [email protected]

Bacillus velezensis G341 isolated from ginseng root suppresses the mycelial growth of various plant pathogens. In order to select optimum solid-state fermentation (SSF) conditions for improving the production ofantifungal metabolites by B. valezensis G341, the bacterium was cultured in different composition solid media consisting of soybean curb residue, zeolite, vermiculite and/or rice bran for 7 days at 30℃. SSF product using soybean curb residue:rice bran:vermiculite (5:2:13, w/w/w) was selected by both in vitro and in vivo antifungal bioassays using Magnaporthe oryzae.The SSF product of G341 was more effective than that of liquid fermentation broth in controlling rice blast. SSF product effectively suppressed the development of phytophthora blight of red pepper causedby Phytophthora capsici, whereas its liquid fermentation broth was not active. Evaluation of the disease control efficacy of the SSF product of G341 is in progress on various soil-borne diseases. The SSF product of B. velezensis G341 could be applied as a microbial fungicide for the controlof plant diseases. Keywords : Bacillus velezensis, solid-state fermentation, antifungal activity

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Comparison Analysis of Secondary Metabolite Profiling BetweenStreptomyces Griseus and its rsmG Mutant Changmin SUNG2, Pyoung Il KIM1, Kozo OCHI3 and Byung-Gee KIM*1

1School of Chemical and Biological Engineering, Institute of Molecular Biology and Genetics, Seoul National University, Seoul 151-744, Republic of Korea.2Interdisciplinary Program of Bioengineering, Seoul National University, Seoul 151-744, Republic of Korea. 3Institute of Technology, Department of Health Science, Miyake 2-1-1, Saeki-ku, Hiroshima, 731-5193, Japan. *Corresponding author: [email protected]

Kozo ochi group generated rsmG mutant of S. griseus that activated the secondary metabolite gene clusters by antibiotics treatment. To compareof secondary metabolite profile, growth conditions of wild- and mutant-type strains in S. griseus were optimized for secondary metabolite production using media transferring from GYM complex media to N-acetylglucosamin based minimal media. Each culture broth was harvested by centrifuge and membrane filteration. To collect secreted secondary metabolites as many as possible, supernatants were extracted two extraction method that organic solvent extraction and ammonium sulfate salting out. Crude extracted samples of wild- and mutant-type wereanalyzed and separated by High-performance liquid chromatography usingC18 column. Secondary metabolite analysis was carried out by ESI-LC-Mass spectrometry using tandem mass profile. This work was supported by National Research Foundation of Korea Grant funded by the Korean Government(20090071169) Keywords : Streptomyces griseus, rsmG mutant, secondary metabolite

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Inhibitory Effects of wild Oenanthe Javanica Extracts Fermentedwith Oligosaccarides on HepG2 cell Proliferation Seun-Ah YANG1, Min-Ju KIM2, Kwang-Hwan JHEE3, Hyung-Dae KIM4, In-Seon LEE1,2, Sam-Pin LEE1,2 and In-Seon LEE*1,2

1The Center for Traditional Microorganism Resources, Keimyung University, Daegu, Korea. 2Dep. of Food Science and Technology, Keimyung University, Daegu, Korea. 3Dept. of Applied Chemistry, Kumoh National Institute of Technology, Gumi, Korea. 4Bissell Chunglog Farm, Daegu, Korea. *Corresponding author: [email protected]

Oenanthe javanica (dropwort) is a medicinal food commonly used to treat diseases such as jaundice, hypertension, and polydipsia in Korea, Japan, and China. In this study, we tested the effects of O. javanica extracts that fermented by steeping with oligosaccaride at room temperature for 1 year (1 yr), and then stored at 4°C for 1 (2 yr) or 2 year (3 yr) on human hepatoma HepG2 cells and human normal liver Chang cell to explore the anticancer properties. The extracts of O. javanica were tested on HepG2 and Chang cells proliferations by MTT assay for 48 h. Our results showed significant concentration-dependent inhibitions of HepG2 proliferation by the three extracts, but no effects on Chang cells viability.Especially, the 1 yr extract showed highest inhibitory effect (49.3% inhibition at 0.1 mg/ml) than 2 yr (40.6%) and 3 yr (38.7%). The inhibitoryeffects on cell morphology changes were also observed. In addition, antioxidative properties of the extracts, including total phenolic and flavonoid contents and radicals scavenging activity were examined. Moreover, we analyzed caffeic acid and ferulic acid, known as main active components, by using HPLC and caffeic acid was detected in the crude extracts about 3-4%, but very little amount of ferulic acid was detected. Keywords : Oenanthe javanica, Dropwort, HepG2 cells

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Streptomyces sp. BCNU 1001 with NF-kB Inhibitory Activity Hye Jung CHOI1, Cheol Soo AHN2, Dong Wan KIM3 and Woo Hong JOO*1

1Dept. of Biology and Interdisciplinary Program in Biotechnology, Changwon National University, Changwon. 2Cho-A Pharm. Co, LTD., Haman-kun. 3Dept. of Microbiology, Changwon National University, Changwon. *Corresponding author: [email protected]

A new actinomycete strain designated as BCNU 1001 was isolated, and cultural, morphological and physiological characters, and 16S rRNA sequences of BCNU 1001 were determined in this study. The 16S rRNA sequence of strain BCNU 1001 was analysed to have high similarity value (99%) with Streptomyces lavendulae. The extracts were obtained by partition chromatography using various solvents and ethylacetate (EA) extract exhibited strong activity against human pathogen fungi and yeasts.Furthermore, EA extract exhibited NF-kB inhibitory activity. The active compounds were isolated from the culture of Streptomyces sp. BCNU 1001 by EA extration, column chromatography and recycling preparative HPLC. These compounds exhibited significant antimicrobial activities against causative agents of Candidiasis and Aspergillosis diseases, Candida albicans and Aspergillus niger, respectively. (Acknowledgments; This research was financially supported by the Korean Ministry of Education,Science Technology (MEST) and Korea Institute for Advancement of Technology (KIAT) through the Human Resource Training Project for Regional Innovation) Keywords : Streptomyces lavendulae, human pathogen fungi, NF-kB

inhibitory activity

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Discovery of Inhibitors for Type III Secretion System Related to the Virulence of Phytophathogenic Gram-negative Bacteria Won-Sik CHOI1, Jae Sun MOON1, Sang Han LEE2, Eun Na LEE1, Sung Eun KIM1 and Sung Uk KIM*1

1Division. of Biosystem Research, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-806, Korea. 2Dept. of Food Science and Technology,Kyungpook National University, Daegu 702-701, Korea. *Corresponding author: [email protected]

The type III secretion system (TTSS) displays a potent virulence mechanism of pathogenic Gram-negative bacteria, such as Salmonella, Shigella, Erwinia, Xanthomonas, and Ralstonia species, which cause diseases in humans and plants by the translocation of effector proteins into the cytosol of host cells. Therefore, TTSS inhibitors that target bacterial virulence without detrimental effect on bacterial growth are usefulantibacterial agents for studies of virulence and potential candidates for drug development. To search and isolate TTSS inhibitors of Xanthomonasaxonopodis from natural resources, two hit compounds were selected fromplant extracts. These compounds showed potent inhibitory activities (IC50= 6-12 μg/ml) against the TTSS of Xanthomonas axonopodis without antibacterial activities against various phytopathogenic bacteria, indicatingthat the inhibition of TTSS-mediated protein secretion by compounds was not due to inhibition of bacterial growth. However, these compounds showed weak disease-control activities in vivo against Xanthomonas oryzae pv. oryzae, the causative agents of bacterial leaf blight of rice. Keywords : Virulence, TTSS, Gram-negative bacteria

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322 www.kormb.or.kr

In Vitro Anti-Methicillin-Resistant Staphylococcus Aureus Activity and Antibiotic Compounds Produced by Pseudomonassp. BCNU 2001Uk Hee YANG1, Hye Jung CHOI1, Cheol Soo AHN2, Yong Kee JEONG3, Dong Wan KIM4 and Woo Hong JOO*1

1Dept. of Microbiology, Changwon National University, Changwon 641-773.2Cho-A Pharm. Co, LTD., Haman-kun 637-810, Korea. 3Dept. of Biotechnology, Dong-A University, Busan 604-714, Korea. 4Dept. of Microbiology, Changwon National University, Changwon 641-773. *Corresponding author: [email protected]

Strain BCNU 2001 was isolated from soil samples collected from Tea-baek Mountain area. The 16S rRNA gene sequences and the substratespectrum of the isolate revealed that the strain belonged to the genus Pseudomonas. The supernatants of this culture expressed antimicrobial activity against various kind of bacteria or fungi. The antibacterial activities against Methicillin-Resistant Staphylococcus aureus (MRSA) were also assessed in vitro. Hexane, dichloromethane, ethylacetate and peptide extracts of BCNU 2001 were determined to have antibacterial principles against various kind of bacteria and MRSA. Compound A was isolated from the culture of Pseudomonas sp. BCNU 2001 by solvent extraction, silica gel column chromatography, HPLC and recycling preparative HPLC. Its structure was elucidated by spectroscopic analyses including mainly various NMR experiment. (This research was financially supported by the Korean Ministry of Education, Science Technology (MEST) and Korea Institute for Advancement of Technology (KIAT) through the Human Resource Training Project for Regional Innovation) Keywords : Tea-baek Mountain, Methicillin-Resistant Staphylococcus

aureus (MRSA), Pseudomonas

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Novel Polyene Macrolides Produced by Recombinant Pseudonocardia autotrophica Strains with Genetically Altered Polyketide Synthase Mijin LEE1, Kyuboem HAN2 and Eung-Soo KIM*1

1Department of Biological Engineering, Inha University, Incheon 402-751, Korea.2CHA BIO & DIOSTECH Bio Pharmaceutical Research Institute, 461-6 Jeonmin-dong, Yuseong, Daejeon 305-811, Korea. *Corresponding author: [email protected]

The polyene antibiotics including nystatin, pimaricin, amphotericin and candicidin comprise a family of very promising antifungal polyketide compounds, and are abundantly produced by streptomycetes and some other soil bacteria. Using the genomics-guided polyene screening method, a rare actinomycetes called Pseudonocardia autotrophica was previously identified to contain a functionally-clustered nystatin-like biosynthetic genes and to produce a presumably novel polyene compound named NPP. Many previous researches revealed that modification of the polyene structure through engineering of its biosynthetic genes is a promising strategy for the production of novel polyene derivatives suitable for potential pharmaceuticals. In our research, we have obtained new derivatives by engineering of genes related with both early and late stages of NPP biosynthetic pathways. This biosynthetic modification approach is already yielding sufficient amount of novel materials for testing the toxicity and bioactivity, thus opening possibilities for discovery of drug-leads for development of effective and safe pharmaceuticals. More detailed examples of P. autotrophica genome engineering techniques and bioactivities of new derivatives will be further discussed. Keywords : polyene, Pseudonocardia autotrophica, Nystatin-like

Pseudonocardia Polyene (NPP)

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Beauvericin, Isolated from the Soil Fungus F00112, a Inhibitor ER Stress Induced XBP-1 Splicing and GRP78 in Fao Cells Jong Won KIM, Eun Young BEA, Mei Hua NAN, Sun Mi LEE, Jae Hyuk JANG, Bo Yeon KIM and Jong Seog AHN*Chemical Biology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Chengwongun 363-883, Korea. *Corresponding author: [email protected]

The endoplasmic reticulum (ER) is a perinuclear, cytoplasmic compartment where proteins and lipids are synthesized. The 78-kDa glucose protein GRP78 is a central regulator of endoplasmic reticulum (ER) homeostasis, functioning in protein folding, ER calcium binding and modulation of transmembrane ER stress sensor activity. ER stress uncouples the interaction between GRP78 and ER stress sensors, leadingto the activation of unfolded protein response (UPR), including upregulation of ER chaperone proteins. In the course of screening for an inhibitor of ER stress-induced XBP-1 splicing and regulators of GRP78/Bipmolecular chaperone expression, we detected potent inhibitory effect in an extract of the soil fungus, F00112. In this sutdy, we discuss isolation,structure identification, and biological activity of cyclohexadepsipeptide type metabolite, beauvericin, as ER stress inhibitor. Keywords : ER stress inhibitor, beauvericin, fungal strain

F-69

G _ Biocatalysis and Fermentation Technology

Isolation and characterization of a novel lipolytic enzyme frommetagenomic library of Ibam mountain in South Korea Soon-Ok RIM1, Bong-Seok SHIN1,2, Yun-Jon HAN1,2, Kyong-Cheol KO1, Jong-Hyun CHOI1, Geun-Joong KIM2 and Jae-Jun SONG*1

1Enzyme Based Fusion Technology Research Team, Microbe-based Fusion Technology Research Center, KRIBB, 1404 Sinjeong-dong, Jeongeup-si, Jeonbuk 580-185, Korea. 2Department of Biological Sciences, College of Natural Sciences,Chonnam National University, 300 Yong-Bong Dong, Buk-Gu, Gwangju 500-757, Republic of Korea. *Corresponding author: [email protected]

Lipolytic enzymes catalyze the hydrolysis and the synthesis many fattyacid esters. The construction of metagenomic library has been used successfully to discovery a wide variety of novel enzyme from environments. A new lipolytic enzyme was isolated from a metagenomic library which was constructed from Korea Ibam mountain,. We screened new lipase/esterase gene that encoded a lipolytic enzyme on 1% tributyrinLB-agar plate. The new lipase/esterase gene had an open reading frame (ORF) of 936 base pairs encoding 311 amino acids. The amino acid sequence was 62% identical to that of uncultured bacterium and belongswith family IV esterase. Our finding of new novel family IV esterase emphasizes the importance of environmental metagenomic libraries as a source of isolation of novel genes and this new esterase will be useful enzyme in this family. (This work was supported by a high throughput screening (HTS)-based Integrated Technology Development grant from the MEST, and a basic research grant from KRIBB.) Keywords : metagenomic library, lipolytic enzymes, p-nitrophenyl-esters

G-1

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