Hsp Proteins

Heat Shock Proteins and Molecular Chaperones

Heat Shock Proteins and Molecular Chaperones Antibodies Proteins ELISA Kits

Hsp10 Hsp27 Hsp32 Hsp40 Hsp60 Hsp70 Hsp90 Hsp110 Chaperones

Heat Shock Proteins and Molecular Chaperones

Hsp10 ....................................3 Hsp27 ....................................3 Hsp32 ....................................5 Hsp40 and the DnaJ Family ...7 Hsp60 and GroEL ..................9 Hsp70 and DnaK.................11 Hsp90 ..................................13 Hsp110 ................................15 Chaperones & Others.........16 Hsp27 Review .....................20 Hsp70 Review .....................22 Hsp90 Review .....................24 References ..........................26Assay Designs is committed to providing scientists with reliable tools for scientific discovery. Our merger in 2005 with Stressgen Bioreagents has broadened our product offering to include an extensive collection of heat shock protein related products including ELISA kits, antibodies, recombinant proteins, and reagents. We are pleased to add the high quality products generated by over 15 years of Stressgen research into our product line, and are dedicated to the further development and manufacturing of novel userfriendly heat shock protein related products. Assay Designs aims to Simplify Your Science by offering this guide to the heat shock protein families, and the products we offer to advance research in this existing field. Heat shock proteins are ubiquitously expressed polypeptides whose expression increases in response to a variety of different metabolic insults. Despite their designation, most of the heat shock proteins are constitutively expressed and perform essential functions. Most notable is their role as molecular chaperones, facilitating the synthesis and folding of proteins throughout the cell. In addition, heat shock proteins have been shown to participate in protein assembly, secretion, trafficking, protein degradation, and the regulation of transcription factors and protein kinases. Increased levels of the heat shock proteins after stress plays a central role in cellular homeostasis.

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Species:H: human M: mouse R: rat B: bovine C: chicken D: drosophila Y: yeast

Bei Biomol erhalten Sie die Assay Design- und Stressgen-Produkte:Bestellen in Deutschland: Biomol GmbH Fon: 0800-246 66 51 Fax: 0800-246 66 52 [email protected] www.biomol.de Technischer Support: [email protected]

Applications:WB: western blot IP: immunoprecipitation ICC: immunocytochemistry IHC: immunohistochemistry F: flow cytometry

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Hsp10Hsp10, also known as Chaperonin 10 (Cpn10), is the ~10 kDa mammalian equivalent of the bacterial GroES gene product. Hsp10 exists in vivo as an oligomer and interacts with Hsp60, the mammalian homolog of the bacterial GroEL protein. Together the Hsp10/Hsp60 chaperonin present within mitochondria facilitates the folding of newly synthesized proteins and may participate in the refolding of proteins damaged after stress. In plants, a similar chaperonin system operates within chloroplasts and is referredEntrez Gene ID3336

Assay Designs (Stressgen) ProductsCat. No. SPA-110 SPA-210 SPP-620 Product Description Hsp10 (Cpn10) Polyclonal Antibody GroES Polyclonal Antibody GroES Recombinant Protein Species H, M, R, X E.coli E.coli Application WB, IP WB, IP

to as the Rubisco-Binding protein. Finally, in bacteria GroEL and GroES participate in the folding and assembly of numerous proteins.Tissue DistributionBroad

Official SymbolHSPE1

NameHeat Shock 10kDa Protein 1 (Chaperonin 10)

SynonymsCPN10, GROES, HSP10

Biological FunctionCaspase activation, Chaperone binding, Protein folding

Cellular DistributionMitochondria Chloroplasts

Hsp27Assay Designs (Stressgen) ProductsHsp27 (sometimes referred to as Hsp20, Hsp25, Hsp28 or the low molecular weight heat shock protein) is homologous to the a Crystallin proteins. Both families of proteins are characterized by their oligomeric structure and are thought to function as ATP-independent chaperones. Hsp27 structure/function is thought to be modulated by phosphorylation mediated by different protein kinases. In addition to their chaperone role, Hsp27 and aB-Crystallin have been shown to mediate structural integrity and membrane stability, affecting actin polymerization, intermediate filament organization, apoptosis, and invasive potential. Evidence suggests altered expression of small heat shock proteins is implicated in the pathogenesis of human diseases including cancer, cataracts, neurodegenerative disorders, and cardiovascular disease.Cat. No. EKS-500 SPA-796 SPA-801 SPA-803 SPA-800 SPA-525 SPA-523 SPA-524 905-642 SPA-800FI SPA-800B SPA-221 SPA-222 SPA-223 SPA-224 SPA-225 SPA-226 SPA-227 SPA-230 NSP-510 SPP-715 ESP-715 SPP-226 SPP-227 SPP-225 Product Description Hsp27 ELISA Kit Hsp20 Polyclonal Antibody Hsp25 Polyclonal Antibody Hsp27 Polyclonal Antibody Hsp27 Monoclonal Antibody (G3.1) Hsp27 (phospho-Ser15) Polyclonal Antibody Hsp27 (phospho-Ser78) Polyclonal Antibody Hsp27 (phospho-Ser82) Polyclonal Antibody Hsp27 (phospho-Ser82) Monoclonal Antibody Hsp27 Monoclonal Antibody, FITC Conjugate Hsp27 Monoclonal Antibody, Biotin Conjugate a A Crystallin Polyclonal Antibody a B Crystallin Monoclonal Antibody a B Crystallin Polyclonal Antibody a A/B Crystallin Polyclonal Antibody

Hsp27 Review on page 20

Species H H, M, R M, R H H, M, R H, M, R H, M, R H, R H, M, R H, M, R H, M, R B H, M, R, B, C H, M, R, B B H, M, B H, M, R, B, C, X M, R, B B M H H B B B

Application

WB WB, IP, ICC, IHC WB, IP, ICC, IHC WB, IP, ICC, IHC WB WB, IP WB, IP WB F, IF

WB, IP WB, ICC, IHC WB, IP WB WB, IP WB, IP WB, IP WB

a B Crystallin (phospho-Ser19) Polyclonal Antibody a B Crystallin (phospho-Ser45) Polyclonal Antibody a B Crystallin (phospho-Ser59) Polyclonal Antibody b Crystallin Monoclonal Antibody (3H92) Hsp25 Recombinant Protein Hsp27 Recombinant Protein Hsp27 Recombinant Protein - Low Endotoxin a A Crystallin Native Protein a B Crystallin Native Protein a A/B Crystallin Native Protein

03

Hsp27 (continued)Official SymbolCRYAA CRYAB

NameCrystallin, a A Crystallin, a B

SynonymsCRYA1, HSPB4 CRYA2, HSPB5

Entrez Gene ID1409 1410

Biological FunctionStructural component of eye, Protein Folding Structural component of eye, Protein Folding, Muscle contraction, Muscle development, Protein folding, Receptor mediated signaling, Visual perception Anti-apoptosis, Cell motility, Protein folding

Tissue DistributionEye Lens Broad

Cellular LocalizationCytoplasm Contractile fibers, Cytoplasm, Nucleus, Plasma membrane Cytoplasm Nucleus, Plasma Membrane

Human DiseasesCataracts Cataracts, Multiple Sclerosis

HSPB1

Heat Shock 27kDa Protein 1

HSP27, Hsp25

3315

Broadly

Charcot-MarieTooth disease, axonal, type 2F. Neuropathy, distal hereditary motor

HSPB2 HSPB3 HSPB6

Heat Shock 27kDa Protein 2 Heat Shock 27kDa Protein 3 Heat Shock Protein, a-Crystallinrelated, B6 Heat Shock 27kDa Protein family, member 7 (cardiovascular) Heat Shock 22kDa Protein 8

MKBP HSPL27 Hsp20

3316 8988 126393

Enzyme Activator, Protein Folding, Somatic Muscle Development Protein Folding Structural component of eye, Protein Folding, Muscle Contraction Protein Folding, Muscle Contraction

Heart, Skeletal Muscle, Skin Muscle Muscle, Ovary

Cytoplasm, Nucleus

Actin cytoskeleton Contractile Fibers

HSPB7

cvHSP

27129

Cardiac, Connective, Skeletal Broadly

HSPB8

H11; E2IG1; HSP22

26353

Kinase Activity, Transferase Activity, Protein Folding

Cytoplasm, Plasma Membrane

CharcotMarie-Tooth disease type 2L. Hereditary motor neuropathy type II

HSPB9

Heat Shock Protein, a-Crystallinrelated, B9 Outer dense fiber of sperm tails 1 HSPB10, ODFP, SODF

94086

Protein Folding

Testes

Nucleus, Cytoplasm Cytoplasm

ODF1

4956

Structural activity, Cell differentiation, Spermatogenesis

Testes

Hsp27 (phospho-Ser82) Polyclonal Antibody (Cat. No. SPA-524)Confocal immunofluorescent staining of Hela cells using Phospho-Hsp (Ser82) Polyclonal Antibody (Cat. No. SPA-524; green) (A). Blue pseudocolor fluorescent dye DRAQ5 was used to stain cell nuclei (B). In (C) the two images are overlapped.

A

B

C

04

H: human, M: mouse, R: rat, B: bovine, C: chicken, D: drosophila, Y: yeast

Hsp32 (Heme Qxygenase)Heme oxygenase or Hsp32 is an esCat. No. Product Description Species Application sential component in the catabolism of heme, catalyzing the first step in EKS-800 HO-1 (Hsp32; Human) ELISA Kit H the degradation of heme to bilirubin. EKS-810 HO-1 (Hsp32; Rat) ELISA Kit R Heme oxygenase consists of at least SPA-895 HO-1 (Hsp32) Polyclonal Antibody H, M, R WB, IP, ICC, IHC, F three isoforms: stress inducible HO-1, SPA-896 HO-1 (Hsp32) Polyclonal Antibody H, M, R WB, IP, ICC, IHC and constitutively expressed HO-2 OSA-150 HO-1 (Hsp32) Polyclonal Antibody H, M, R WB, IP and HO-3. Induction of HO-1 occurs OSA-110 HO-1 (Hsp32) Monoclonal Antibody (HO-1-1) H, M, R WB, F in response to heat shock, oxidative OSA-111 HO-1 (Hsp32) Monoclonal Antibody (HO-1-2) H, M, R WB, IHC, F stress, thiol reacting reagents, heavy OSA-111FI HO-1 (Hsp32) Monoclonal Antibody-FITC Conjugate H, M, R F, IF metals, inflammatory mediators and OSA-111B HO-1 (Hsp32) Monoclonal Antibody-Biotin Conjugate H, M, R certain growth factors. The majoOSA-200 HO-2 Polyclonal Antibody H, M, R WB rity of the protein is localized to SPA-897 HO-2 Polyclonal Antibody H, M, R, C WB, IP, ICC the endoplasmic reticulum, but the SPP-730 HO-1 (Hsp32) Recombinant Protein R protein can also be present at the plasma membrane and mitochondSPP-732 HO-1 (Hsp32) Recombinant Protein H ria. The products produced by heme NSP-550 HO-2 Recombinant Protein H oxygenase have important physiological effects: carbon monoxide is a potent vasodilator; biliverdin and its product bilirubin are antioxidants; and the released iron can increase oxidative stress if not effectively reutilized. Modulation of HO expression may be useful for protection against atherosclerotic disease, oxidative stress, coronary ischemia, hypertension and certain neurodegenerative diseases.

Official SymbolHMOX1 HMOX2

NameHeme Oxygenase 1 Heme Oxygenase 2

SynonymsHO-1 HO-2

Entrez Gene ID3162 3163

Biological FunctionBreakdown of heme, protection against oxidative stress, Ion binding, NFkB signaling Breakdown of heme, protection against oxidative stress, Electron carrier, Ion binding

Tissue DistributionBroad Broad

Cellular DistributionEndoplasmic reticulum Endoplasmic reticulum

Heme-Oxygenase-1 (Hsp32) Monoclonal Antibody (Cat. No. OSA-111)Human lung carcinoma tissue was immunohistochemically stained using Heme-Oxygenase-1 Monoclonal Antibody (clone HO-1-2) at a dilution of 1:50. Another Heme-Oxygenase-1 monoclonal (clone HO-1-1, Cat. No. OSA-110) (1:50) was negative for staining the same lung cancer tissue.

WB: western blot, IP: immunoprecipitation, ICC: immunocytochemistry, IHC: immunohistochemistry, F: flow cytometry

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Hsp32 (continued)

Heme Oxygenase 1 (Hsp32)Crystal structure of human Heme Oxygenase-1 in complex with its substrate Heme (Schuller, D.J. et al. 2002).

Heme-Oxygenase-1 (Hsp32) Monoclonal Antibody (Cat. No. OSA-110)Human lung cancer A2 cells were analyzed by flow cytometry using isotype control antibody (left) or Heme-Oxygenase-1 Monoclonal Antibody (clone HO-1-1; right) at a final concentration of 10g/mL.

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Hsp40 and the DnaJ FamilyMammalian Hsp40, present within the cytosol, is one Assay Designs (Stressgen) Products of a number of family members closely related to Cat. No. Product Description Species Applications the DnaJ protein first described in E.coli. Hsp40/DnaJ SPA-400 Hsp40 Polyclonal Antibody H, M, R, C, X WB, IP, ICC, IHC family members work in concert with the Hsp70/DnaK SPA-450 Hsp40 Monoclonal Antibody (2E1) H, M, R WB, IP proteins, facilitating the hydrolysis of ATP to ADP and SPA-410 DnaJ Polyclonal Antibody E. coli WB, IP thereby helping to lock in the binding of the Hsp70 SPP-400 Hsp40 Recombinant Protein H chaperone to its protein substrate. Consequently, SPP-640 Active DnaJ Recombinant Protein E. coli each of the different members of the Hsp70 family in eukaryotes require a specific DnaJ homolog for their chaperone activity. More than 20 genes encoding DnaJrelated proteins have been identified in yeast. In animal cells the Hsp40 family of proteins display a broad tissue distribution with individual members present within most intracellular compartments. Assay Designs currently provides both the DnaJ and Hsp40 proteins in purified form, along with a number of antibodies specific for each protein. New products targeting other members of the DnaJ/Hsp40 family are expected in the near future.

Official SymbolDNAJA1

NameDnaJ (Hsp40) homolog, subfamily A, member 1

SynonymsDJ-2; DjA1; HDJ2; HSDJ; HSJ2; HSPF4; hDJ-2

Entrez Gene ID3301

Biological FunctionProtein binding, Protein folding, LDL receptor binding, Ion binding

Tissue DistributionBroadly

Cellular DistributionCytoplasm Nucleus Nucleolus Golgi Cytoplasm Nucleus Mitochondrion Mitochondrion

DNAJA2

DnaJ (Hsp40) homolog, subfamily A, member 2 DnaJ (Hsp40) homolog, subfamily A, member 3 DnaJ (Hsp40) homolog, subfamily A, member 4 DnaJ homology subfamily A, member 5 DnaJ (Hsp40) homolog, subfamily B, member 1 DnaJ (Hsp40) homolog, subfamily B, member 2 DnaJ (Hsp40) homolog, subfamily B, member 4 DnaJ (Hsp40) homolog, subfamily B, member 5 DnaJ (Hsp40) homolog, subfamily B, member 6 DnaJ (Hsp40) homolog, subfamily B, member 7 DnaJ (Hsp40) homolog, subfamily B, member 8 DnaJ (Hsp40) homolog, subfamily B, member 9

CPR3; DJA2; DNAJ; DNJ3; RDJ2; HIRIP4 TID1, hTid-1

10294

Protein binding, Ion binding, Cell cycle, Protein Folding GTPase activity, Ion Binding, GPCR Signaling, Protein folding, Protein folding, Regulation of apoptosis Protein binding, Ion binding, Protein folding Protein binding, Nucleic acid binding, Ion binding, Protein folding Protein binding, Protein Folding Protein binding, Protein folding Protein binding, Protein folding Protein binding, Protein folding Protein binding, Protein folding Protein Binding, Protein folding Protein binding, Protein folding Chaperone, Protein folding, activity, Protein binding

Broadly

DNAJA3

9093

Broad

DNAJA4 DNAJA5 DNAJB1 DNAJB2 DNAJB4 DNAJB5 DNAJB6 DNAJB7 DNAJB8 DNAJB9

MST104; MSTP104; PRO1472 GS3 protein Hdj1; HSPF1; Hsp40 HSJ1; HSPF3 DjB4; HLJ1; DNAJW Hsc40 MRJ; HSJ2; HHDJ1; HSJ-2; MSJ-1 HSC3 MGC33884 MDG1; ERdj4

55466 134218 3337 3300 11080 25822 10049 150353 165721 4189

Broad Broad Broad Broad Broad Broad Broad Broad Cytoplasm Nucleus Cytoplasm Nucleus Nucleus Cytoplasm Nucleus

Broad

Cytoplasm Endoplasmic reticulum Nucleolus Nucleus Cytoplasm Endoplasmic reticulum Nucleus

DNAJB11

DnaJ (Hsp40) homolog, subfamily B, member 11

EDJ; ERj3; HEDJ; hDj9; ABBP2; ERdj3; ABBP-2

51726

Protein binding, Protein folding

Broad

datasheet: www.biomol.de www.antibodyworld.com

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Hsp40 (continued)Official Symbol DNAJB12 DNAJB13 DNAJB14 DNAJC1 Name DnaJ (Hsp40) homolog, subfamily B, member 12 DnaJ (Hsp40) related, subfamily B, member 13 DnaJ (Hsp40) homolog, subfamily B, member 14 DnaJ (Hsp40) homolog, subfamily C, member 1 HTJ1; ERdj1; DNAJL1 Synonyms DJ10 TSARG5; TSARG6 Entrez Gene ID 54788 374407 79982 64215 Biological Function Protein binding, Protein folding Protein binding, Apoptosis, Protein folding, Spermatogenesis Protein binding, Protein folding ATPase activation, Protein folding, Chaperone folding, DNA binding Endoplasmic reticulum Membrane Microsome Nucleus Nucleus Testis Tissue Distribution Cellular Distribution Plasma membrane

DNAJC2

DnaJ (Hsp40) homolog, subfamily C, member 2 DnaJ (Hsp40) homolog, subfamily C, member 3 DnaJ (Hsp40) homolog, subfamily C, member 4 DnaJ (Hsp40) homolog, subfamily C, member 5 DnaJ (Hsp40) homolog, subfamily C, member 5 b DnaJ (Hsp40) homolog, subfamily C, member 5 g DnaJ (Hsp40) homolog, subfamily C, member 6 DnaJ (Hsp40) homolog, subfamily C, member 7 DnaJ (Hsp40) homolog, subfamily C, member 8 DnaJ (Hsp40) homolog, subfamily C, member 9 DnaJ (Hsp40) homolog, subfamily C, member 10 DnaJ (Hsp40) homolog, subfamily C, member 11 DnaJ (Hsp40) homolog, subfamily C, member 12 DnaJ (Hsp40) homolog, subfamily C, member 13 DnaJ (Hsp40) homolog, subfamily C, member 14 DnaJ (Hsp40) homolog, subfamily C, member 15 DnaJ (Hsp40) homolog, subfamily C, member 16 DnaJ (Hsp40) homolog, subfamily C, member 17 DnaJ (Hsp40) homolog, subfamily C, member 18 DnaJ (Hsp40) homolog, subfamily C, member 19 HscB iron-sulfur cluster co-chaperone homolog (E. coli) HscB iron-sulfur cluster co-chaperone homolog (E. coli)

Zrf1; Zrf2; MIDA1

22791

DNA binding, Cell cycle, Protein binding, DNA replication, Protein folding, Transcription Protein binding, Protein folding, Kinase inhibitor, Defense Protein binding, Protein folding Protein binding, Protein folding Protein binding, Protein folding Protein binding, Protein folding Protein binding, Signal transduction, Hydrolase activity, Protein folding, Phosphatase activity Protein binding, Protein folding Protein binding, Protein folding Protein binding, Protein folding Protein binding, Redox homeostasis, Protein folding Protein binding, Protein folding Protein binding , Protein folding Protein binding, Protein folding Protein binding, Protein folding Protein binding, Protein folding Protein binding, Protein folding RNA binding, Protein folding, Protein binding Protein binding, Protein folding Protein binding, Protein folding, Protein transport Chaperone binding, Protein binding, Protein folding Broad Broad Broad Broad Broad Broad Broad Broad Broad Pituitary Gland Broad

DNAJC3 DNAJC4 DNAJC5 DNAJC5B DNAJC5G DNAJC6

P58; HP58; PRKRI; P58IPK HSPF2; MCG18; DANJC4 CSP CSP-b CSP-g DJC6

5611 3338 80331 85479 285126 9829

Cytoplasm Membrane Membrane

Membrane Nucleus

DNAJC7 DNAJC8 DNAJC9 DNAJC10 DNAJC11 DNAJC12 DNAJC13 DNAJC14 DNAJC15 DNAJC16 DNAJC17 DNAJC18 DNAJC19 DNAJC20

TPR2; TTC2; DANJC7 SPF31; HSPC331 JDD1; SB73 JPDI; ERdj5

7266 22826 23234 54431 55735

Nucleolus

Endoplasmic Reticulum Mitochondrion

JDP1 RME8 DNAJ; HDJ3; LIP6 MCJ; HSD18; DNAJD1

56521 23317 85406 29103 23341 55192 202052

Endosome Endoplasmic reticulum

Membrane

Membrane Membrane Mitochondrion Mitochondrion

TIM14; TIMM14 HSCB, JAC1; HSC20

131118 150274

HSCB

DNAJC20

150274

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Hsp47 Monoclonal Antibody (Cat. No. SPA-470)Human breast cancer tissue was immunohistochemically stained using Hsp47 Monoclonal Antibody (clone M16.10A1) at a dilution of 1:50.

Hsp60 and GroEL: the ChaperoninsMembers of the Hsp60 (eukaryotes) and GroEL (bacterial) family of heat shock proteins are some of the best characterized molecular chaperones. Bacterial GroEL, named because of its essential role in bacteriophage growth, exists as a large homo-oligomeric complex which recognizes and binds to unfolded polypeptides. In combination with its particular co-factor (Hsp10 in eukaryotes, GroES in bacteria) the Hsp60/ GroEL proteins bind newly synthesized polypeptides and facilitate their folding to the native state via one or more rounds of ATP hydrolysis. Mammalian Hsp60 is localized within mitochondria, while a related form of the protein termed Rubisco-binding protein operates within plant chloroplasts. Finally, within the eukaryotic cytosol, related proteins referred to as CCT or TRIC make a hetero-oligomeric structure and have been shown to similarly bind select protein substrates and facilitate their folding and/or higher order assembly. Oftentimes the GroEL/ES or Hsp60/Hsp10 protein folding machineries are referred to as the chaperonins. In vitro as well as in vivo chaperonins, either alone or with other chaperones and ATP, have been shown to orchestrate the re-folding of partially denatured proteins. In addition to their prominent role as molecular chaperones members of the GroEL and Hsp60 families have long been recognized as

Assay Designs (Stressgen) ProductsCat. No. EKS-600 EKS-650 SPA-807 SPA-806 SPA-829 SPA-828 SPA-881 SPS-870 SPS-875 SPA-110 SPA-210 CTA-123 CTA-191 CTA-202 ESP-540 Product Description Hsp60 ELISA Kit (measures protein) Hsp60 ELISA Kit (measures antibodies) Hsp60 Monoclonal Antibody (LK-2) Hsp60 Monoclonal Antibody (LK-1) Hsp60 Monoclonal Antibody (11-13) Hsp60 Goat Polyclonal Antibody Hsp65 Monoclonal Antibody (3F7) GroEL Monoclonal Antibody (9A1/2) GroEL Polyclonal Antibody Hsp10 (Cpn10) Polyclonal Antibody GroES Polyclonal Antibody TCP-1a Monoclonal Antibody (23c) TCP-1a Monoclonal Antibody (91a) TCP-1b Monoclonal Antibody Active Hsp60 Recombinant Protein (Low Endotoxin) NSP-540 ESP-741 Active Human Hsp60 Recombinant Protein Active Mouse Hsp60 Recombinant Protein (Low Endotoxin) SPP-741 SPP-742 NSP-581 SPP-610 SPP-620 Hsp60 Recombinant Protein Hsp60 Recombinant Protein Hsp65 Recombinant Protein Active GroEL Recombinant Protein GroES Recombinant Protein M R Mycobacterim E. coli E. coli H M Species H H H, M, R, C, Y H, M, R, C, X H, M, R, D H, M, R, C, X H, M, R, D E. coli H, M, R, Y, E. coli H,,M, R, X E. coli M, R H, M, R, D, Y H, M, R, C H WB, IHC, F WB, IP, F WB, IP, IHC, F WB, IP WB WB, IP WB W, IP W, IP W, IP, ICC WB, IP, F WB Applications


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Hsp60 (continued)highly immunogenic proteins and consequently have attracted much attention from immunologists. Assay Designs offers a comprehensive panel of both purified chaperonin proteins isolated from different sources along with antibodies capable of discerning the different family members.

Official SymbolHSPD1

NameHeat Shock 60kDa Protein 1 (chaperonin)

SynonymsCPN60; GROEL; HSP60; HSP65; SPG13; HuCHA60

Entrez Gene ID3329

Biological FunctionProtein folding, Nucleotide binding, Protein import, Regulation of apoptosis

Tissue DistributionBroad

Cellular DistributionMitochondrion Cytoplasm

Human DiseasesSpastic paraplegia

Hsp60 Monoclonal Antibody (Cat. No. SPA-807)Human colon cancer tissue was immunohistochemically stained using Hsp60 Monoclonal Antibody (clone LK-2) at a dilution of 1:50.

Hsp60 Monoclonal Antibody (Cat. No. SPA-829)Human hepatoma tissue was immunohistochemically stained using Hsp60 Monoclonal Antibody (clone LK-1) at a dilution of 1:50. Another Hsp60 monoclonal (clone 11-13, Cat. No. SPA-806) (1:50) was negative for staining the same hepatoma tissue.

Hsp60 Monoclonal Antibody (Cat. No. SPA-806)Human hepatoma QGY cells were analyzed by flow cytometry using isotype control antibody (left) or Hsp60 Monoclonal Antibody (clone LK-1; right) at a final concentration of 10g/mL.

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Hsp70 and DnaKThe Hsp70 family of heat shock proteins contains multiple homologues ranging in size from 66kDa to 78kDa. These proteins include cognate members which are found within major intracellular compartments, and highly inducible isoforms which are predominantly cytoplasmic or nuclear in distribution. All Hsp70 family members contain a highly conserved N-terminal ATPase domain, as well as a conserved hydrophobic peptide binding domain (PBD) and more variable a-helical cap domain. Activation of Hsp70 is coordinated by binding of ATP at the N-terminus, causing a conformational change that opens the cap, allowing interaction of the PBD with a wide variety of client proteins in their unfolded, misfolded, or denatured state. Hsp70 ATPase activity is influenced by association with

Hsp70 Review on page 22specific co-chaperone molecules, including Hsp40, Hip, Hop, Hup, Hap, and CHIP. These co-chaperones cooperate with Hsp70 to fold newly synthesized proteins, re-fold misfolded or denatured proteins, coordinate trafficking of proteins across cellular membranes, disassemble clathrin-coated vesicles, inhibit protein aggregation, and target the degradation of proteins via the proteasomal pathway. Elevated levels of Hsp70 have been associated with inhibition of apoptosis, and clinical correlations between Hsp70 expression and poor cellular differentiation, increased lymph node metastasis, chemoresistance, and poor clinical outcome indicate Hsp70 may be of use in the diagnosis, prognosis, and treatment of human malignancy.

Assay Designs (Stressgen) ProductsCat. No. EKS-700 EKS-750 EKS-725 SPA-810 SPA-810FI SPA-810B SPA-812 SPA-811 SPA-820 SPA-822 SPA-757 SPA-815 SPA-816 SPA-756 SPA-754 SPA-766 SRA-1500 SPS-825 SPA-826 NSP-555 ESP-555 SPP-758 SPP-751 SPP-752 Product Description Hsp70 ELISA Kit (assay protein) Hsp70 ELISA Kit (assay antibodies) Hsp70B ELISA Kit (assay protein) Hsp70 (Hsp72) Monoclonal Antibody (C92F3A-5) Hsp70 (Hsp72) Monoclonal Antibody, FITC Conjugate Hsp70 (Hsp72) Monoclonal Antibody, Biotin Conjugate Hsp70 (Hsp72) Polyclonal Antibody Hsp70 (Hsp72) Polyclonal Antibody Hsp70/Hsc70 Monoclonal Antibody (N27F3-4) Hsp70/Hsc70 Monoclonal Antibody (BB70) Hsp70/Hsc70 Polyclonal Antibody Hsc70 (Hsp73) Rat Monoclonal Antibody (1B5) Hsc70 (Hsp73) Polyclonal Antibody Hsp70B Polyclonal Antibody Hsp70B Monoclonal Antibody (175f) Hip Polyclonal Antibody HOP (p60) Monoclonal Antibody (DS14F5) Grp75 Monoclonal Antibody (30A5) Grp78 (BiP) Polyclonal Antibody Active Hsp70 (Hsp72) Recombinant Protein Active Hsp70 Recombinant Protein (Low Endotoxin) Active Hsp70 (Hsp72) Recombinant Protein Active Hsc70 (Hsp73) Recombinant Protein Active Hsc70 (Hsp73) Recombinant Protein, ATPase fragment ESP-502 Active Hsp70-A2 Recombinant Protein (Low Endotoxin) SPP-762 SPP-767 SRP-1510 SPP-765 Hsp70B Recombinant Protein Hip Recombinant Protein HOP (p60) Recombinant Protein Grp78 (BiP) Recombinant Protein H R H Hamster M Species H, M, R H H H, M, R, C, D H, M, R, C, D H, M, R, C, D H, M, R, F H, M, R H, M, R, C, X H, M, R, C, X H, M, R, C, Y H, M, R, C H, M, R H H H, M, R, B H, M, R, C, X H, M, R, C, D, X M, R, X H H R B B WB, IP, ICC, IHC WB WB, F WB, IHC, F WB, IP WB, IP, ICC, IHC, F WB, ICC, IHC WB, IP WB WB WB, IP WB, IHC WB, IP, ICC, IHC WB, IHC, F Applications


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Hsp70 (continued)Official Symbol HSPA1A Name Heat Shock 70kDa Protein 1A Synonyms HSP72, HSPA1, HSPA1B, HSP70-1 Entrez Gene ID 3303 Biological Function Nucleotide binding, Protein folding Tissue Distribution Broadly Cellular Distribution Cytoplasm, Nucleolus Human Diseases Alzheimers, Ankylosing spondylitis, Asthma, Chronic obstructive pulmonary , Multiple sclerosis, Parkinsons disease, Restenosis, Schizophrenia, Tuberculosis Asthma, Alzheimers, Obesity, Chronic obstructive pulmonary, Crohns disease, Diabetes (type 1 & 2), Hypothyroidism, Multiple, Pancreatitis, Parkinsons disease, Schizophrenia, Sclerosis, Restenosis Chronic obstructive pulmonary disease, Graft vs. Host disease, Hypothyroidism, Multiple sclerosis, Parkinsons disease, Restenosis, Schizophrenia, Septic shock, Storke Alzheimers, Crohns, Sepsis

HSPA1B

Heat Shock 70kDa Protein 1B

HSP70-2

3304

Nucleotide binding, Protien folding, Regulation of apoptosis

Broad?

Cytoplasm, Endoplasmic reticulum, Mitochondrion Nucleus Cytoplasm, Nucleus

HSPA1L

Heat Shock 70kDa Protein 1-like

Hum70t, HSP70-HOM

3305

Nucleotide binding, DNA repair, Protein folding, Telomere maintenance

Broad?

HSPA2

Heat Shock 70kDa Protein 2 Heat Shock 70kDa Protein 4 RY, APG2, Hsp70, Hsp70RY, HS24/P52 BIP, MIF2, GRP78

3306

Nucleotide binding, Protein folding, Spermatogenesis Nucleotide binding, Protein folding

Broad

Nucleus, Cytoplasm, Nucleolus Cytoplasm

HSPA4

3308

Broad

Preterm delivery, Carotid Plaques

HSPA5

Heat Shock 70kDa Protein 5 (glucose-regulated protein, 78kDa) Heat Shock 70kDa Protein 6 (HSP70B) Heat Shock 70kDa Protein 7 (HSP70B) Heat Shock 70kDa Protein 8

3309

Nucleotide binding, Apoptosis, Caspase regulation, Protein binding, Ribosome binding

Broad

Endoplasmic Reticulum, Plasma Membrane, Cytoplasm, Nucleolus, Nucleus

Bipolar disease, Schizophrenia

HSPA6

3310

Nucleotide binding, Protein folding Nucleotide binding, rotein folding Nucleotide binding, Protein folding, ATPase activity Broad

HSPA7

HSP70B

3311

HSPA8

LAP1, HSC54, HSC70, HSC71, HSP71, HSP73, NIP71, HSPA10 CSA, MOT, MOT2, GRP75, HSPA9, PBP74, Mot-2 HSP70-4; HSP70L1

3312

Cytoplasm, Nucleus

Cystic Fibrosis, Lung cancer

HSPA9

Heat Shock 70kDa Protein 9 (Mortalin) Heat Shock 70kDa Protein 14

3313

Nucleotide binding, Protein folding, Regulation of apoptosis

Cytoplasm Mitochondria

HSPA14

51182

Hsp70Substrate binding domain of Hsp70 in complex with a substrate peptide. Science (1996) 272(5268):1606-1614.

Hsp70 Monoclonal Antibody (Cat. No. SPA-810)Human colon cancer tissue was immunohistochemically stained using Hsp70 Monoclonal Antibody (clone C92F3A-5) at a dilution of 1:50.

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Hsp90The 90kDa molecular chaperone family comprises several proteins including the 90 kDa heat shock protein Hsp90 and the 94kDa glucose-regulated protein grp94, which are major molecular chaperones of the cytosol and endoplasmic reticulum. In mammalian cells there are at least two Hsp90 isoforms, Hsp90a and Hsp90b, which are encoded by separate genes. All known members of the Hsp90 protein family are highly conserved, especially in the N-terminal and C-terminal regions which contain independent chaperone sites with different client protein specificity. Hsp90 is part of the cells network of chaperones that regulate protein folding and assembly, requiring both ATP and co-chaperones (e.g. Hsp70, Hsp40, Hip/Hop, p23, and Aha1) for function. Inhibition of the Hsp90 protein folding

Hsp90 Review on page 24machinery targets client proteins for ubiquitin-mediated proteolysis. Hsp90 is also a necessary component of fundamental cellular processes such as hormone signaling, cell growth, and differentiation through its binding of client proteins such as ErbB2/Her-2, Akt, Raf, CDK1, and CDK4. In addition to its homeostatic and stress induced roles in protein folding, grp94 can function in the intracellular trafficking of peptides from the extracellular space to the MHC class I antigen processing pathway of antigen presenting cells. Strategies seeking to disrupt Hsp90 activation of key pro-survival pathways hold promise as either direct or adjuvant therapies for cancers displaying altered Hsp90 expression.

Assay Designs (Stressgen) ProductsCat. No. EKS-895 SPA-830 SPA-835 SPA-845 SPA-846 SPS-771 SPA-840 SPA-843 SPA-842 SRA-1400 SPA-670 SPP-776 SPP-770 SPP-670 HPK-102 HPK-101 Product Description Hsp90a ELISA kit Hsp90 Monoclonal Antibody (AC88) Hsp90 Rat Monoclonal Antibody (16F1) Hsp90 Rat Monoclonal Antibody (2D12) Hsp90 Polyclonal Antibody Hsp90a Polyclonal Antibody Hsp90a Rat Monoclonal Antibody (9D2) Hsp90b Monoclonal Antibody (K3701) Hsp90b Monoclonal Antibody (K3705) FKBP-59 (Hsp56, p59) Monoclonal Antibody p23 Polyclonal Antibody Hsp90a Recombinant Protein Hsp90 Native Protein p23 Recombinant Protein Geldanamycin 17-AAG Species H H, M, R, C H, M, R, C, D H, M, R, C H, M, R, C H, M, R, C, X H, C H, M, R H, M, R, C H, M, R H, M, R H H H WB, IP, ICC, IHC, F WB, IP, ICC, IHC WB, IP WB WB, IP WB, IHC, F WB, F WB, IHC WB, IP, ICC, IHC WB Applications

Official Symbol HSP90AA1

Name Heat Shock Protein 90kDa a (cytosolic), class A member

Synonyms HSPN; LAP2; HSP86; HSPC1; HSPCA; Hsp89; Hsp90; HSP90A; HSP90N; HSPCAL1; HSPCAL4 HSP90ALPHA, HSPCA, HSPCAL3 HSPC2; HSPCB; D6S182; HSP90B; FLJ26984; HSP90-BETA GP96; GRP94

Entrez Gene ID 3320

Biological Function Nucleotide binding, Protein folding, Protein dimerization, Signal transduction

Tissue Distribution Broadly

Cellular Distribution Cytoplasm, Nucleus

Human Diseases

HSP90AA2

Heat Shock Protein 90kDa a (cytosolic), class A member 2 Heat Shock Protein 90kDa a (cytosolic), class B member 1 Heat Shock Protein 90kDa b (Grp94), member 1

3324

Nucleotide binding, Protein folding Nucleotide binding, Protein folding Ion binding, Anti-apoptosis, Nucleotide binding, Ion sequestration , Protein folding, Protein transport Broad Cytoplasm

HSP90AB1

3326

HSP90B1

7184

Broad

Endoplasmic reticulum

Ischemic neuronal cell death


13

Hsp90 (continued)Hsp90Crystal structure of an Hsp90-Sba1 closed chaperone complex. Nature (2006) 440(7087):1013-1017.

Hsp90a Monoclonal Antibody (Cat. No. SPA-840)Human colon cancer tissue was immunohistochemically stained using Hsp90a monoclonal antibody (clone 9D2) at a dilution of 1:50.

Hsp90b Monoclonal Antibody (Cat. No. SPA-842)Human breast cancer tissue was immunohistochemically stained using Hsp90b monoclonal antibody (clone K3705) at a dilution of 1:50.

14

Hsp90a Monoclonal Antibody (Cat. No. SPA-840)Human colon cancer Coca-2 cells were analyzed by flow cytometry using isotype control antibody (left) or Hsp90a monoclonal antibody (clone 9D2; right) at a final concentration of 10g/mL.

Hsp110Hsp110 belongs to a family of large stress proteins referred to as the Hsp110/SSE (yeast stress seventy) family. Mammalian Hsp110 shares approximately 30% amino acid identity with its distant relative Hsp70, primarily in the conserved ATP-binding domain. Hsp110 is one of the three or four most abundant Hsps in mammalian tissue, with the highest constitutive expression in the brain. Functionally, Hsp110 appears to complex with other chaperones (predominantly Hsp70 and Hsp25) to maintain and repair protein folding. Hsp110 is more efficient than Hsp70 in conferring heat resistance, and has been shown to possess RNA-binding properties via the N-terminal ATPbinding domain. Due to the inherent efficiency of Hsp110 in binding peptide, the molecule has been utilized extensively as an immunoadjuvant to deliver known tumor antigens to antigen presenting cells, generating antigen-specific innate and adaptive anti-tumor responses.

Assay Designs (Stressgen) ProductsCat. No. SPA-1101 SPA-1103 Product Description Hsp110 Polyclonal Antibody Hsp110 Polyclonal Antibody Species H, M, R, B, Y H, M, R, B, X Applications WB, IP WB

Official Symbol HSPH1

Name Heat Shock 105kDa/ 110kDa Protein 1

Synonyms HSP105, HSP105A, HSP105B

Entrez Gene ID 10808

Biological Function Nucleotide binding, Protein binding

Tissue Distribution Protein folding

Cellular Distribution Broad

Human Diseases Cytoplasm


15

Chaperones & OthersThe homeostatic process of protein folding and the protective functions of the protein folding machinery in stress-induced conditions (i.e. heat, starvation, oxidation) are dependent on heat shock proteins, as well as a diverse group of co-chaperones and transcriptional regulators. These molecules include the Hsp70 co-chaperones HIP and HOP, the ER resident chaperones Calnexin and Calreticulin, and Protein DisulfideIsomerase (PDI). ER-resident chaperones are folded in the ER, and retained via their KDEL peptide motif which is bound by the KDEL receptor (Erd2p). Together with the Hsps, these molecules participate in proper glycoprotein folding, prevent premature oligomerization of nascent proteins, regulate Hsp co-chaperone substrate specificity, and promote the rearrangement of disulphide bonds in the secretory pathway.

Assay Designs (Stressgen) ProductsCat. No. SPA-860 SPA-865 SPA-600 SPA-601 VAP-SV003 SPA-585 SPA-725 SPS-720 SRA-1400 SPA-240 SPA-766 SRA-1500 SPA-950 SPA-901 SPA-960 SPA-470 SPA-1040 SPA-827 VAA-PT048 VAM-PT046 SPA-670 SPA-890 SPA-891 VAM-SV021 CTA-123 CTA-191 CTA-202 VAP-PT068 SPP-767 SRP-1510 SPP-900 NSP-535 SPP-650 SPP-670 LYC-HL101 Product Description Calnexin Polyclonal Antibody Calnexin Polyclonal Antibody Calreticulin Polyclonal Antibody Calreticulin Monoclonal Antibody CSP Polyclonal Antibody ERp57 (Grp58) Polyclonal Antibody ERp57 (Grp58) Monoclonal Antibody ERp72 Polyclonal Antibody FKBP59 (Hsp56, p59) Monoclonal Antibody GrpE Polyclonal Antibody Hip Polyclonal Antibody HOP (p60) Monoclonal Antibody HSF-1 Rat Monoclonal Antibody HSF-1 Polyclonal Antibody HSF-2 Rat Monoclonal Antibody Hsp47 (Colligin) Monoclonal Antibody Hsp104 Polyclonal Antibody KDEL Antibody (Grp78, Grp94) Antibody KDEL Receptor Monoclonal Antibody Membrin Monoclonal Antibody p23 Polyclonal Antibody PDI Polyclonal Antibody PDI Monoclonal Antibody Sec6 Monoclonal Antibody TCP-1a Rat Monoclonal Antibody TCP-1a Rat Monoclonal Antibody TCP-1b Rat Monoclonal Antibody UGGT Polyclonal Antibody Hip Recombinant Protein HOP (p60) Recombinant Protein HSF-1 Recombinant Protein Hsp47 (Colligin) Recombinant Protein Active GrpE Recombinant Protein p23 Recombinant Protein HeLa Cell Lysate (Heat Shocked) Species H, M, R H, M, R, C, X H, M, R H M, R, B, C, X H, M, R, B H H, M, R, B H, B E. coli H, M, R, B H, M, R, B, C, X H, M, R, B H, M, R, B, C, D, X H, M, R, B H, M, R, B Y H, M, R, B, C, D, X H, M, R, B, C, D, X H, R, C H, M, R H, M, R, B, X H, M, R, B, C, X H, M, R, B, C M, R, B H, M, R, B, D, Y H, M, R, B, C M, R R H H H E. coli H Application WB WB, IP, ICC WB, IP, ICC, IHC WB, IP, ICC, IHC, F WB, IP, IHC WB WB, IP, IHC WB WB, IP, ICC, IHC WB, IP WB W,B IP WB, IP, ICC WB, IP, ICC WB, IP WB, IHC WB, IP WB, IP, ICC, IHC, F WB, IP, ICC WB, IP, ICC WB WB, IP, ICC WB, ICC, IHC WB, IP, ICC WB, IP, ICC WB, IP, F WB WB

16


Official Symbol AHSA1

Name AHA1, activator of Heat Shock 90kDa Protein ATPase homolog 1 (yeast) chaperone, ABC1 activity of bc1 complex homolog (S. pombe) Calreticulin

Synonyms AHA1, C14orf3, p38 ADCK3, COQ8

Entrez Gene ID 10598

Biological Function ATPase activity, Protein folding, Chaperone activity, Protein binding Kinase activity, Protein folding, Nucleotide binding DNA binding, Calcium homeostasis, Ion binding, Actin organization, Sugar binding, Protein export, Protein binding, Protein folding, Regulation of apoptosis, meiosis and transcription Ion binding, Angiogenesis, Sugar binding, Protein folding, Protein secretion Protein binding, Protein folding, CDK activity Endopeptidase activity, Proteolysis, Peptidase activity Ion binding, Protein folding, Nucleotide binding, Protein transport, Protein binding Exocytosis, Protein transport Isomerase activity, Protein folding, Protein binding

Tissue Distribution Broad

Cellular Distribution Cytoplasm, Endoplasmic Reticulum Mitochondrion

Human Diseases

CABC1

56997

CALR

RO, SSA, cC1qR

811

Broad

Cytoplasm Endoplasmic reticulum Extracellular environment Cytoplasm Endoplasmic reticulum Membrane Cytoplasm

CANX

Calnexin

CNX, IP90, P90

821

Broad

CDC37 ClpP

cell division cycle 37 homolog (S. cerevisiae) ClpP caseinolytic peptidase, ATP-dependent, proteolytic subunit homolog (E. coli) ClpX caseinolytic peptidase X homolog (E. coli) exocyst complex component 3 [Homo sapiens] FK506 binding protein 1A, 12kDa

P50CDC37

11140 8192

Broad

Mitochondrion

ClpX

10845

Broad

Mitochondrion

EXOC3 FKBP1A

SEC6, SEC6L1, Sec6p FKBP1; PKC12; PKCI2; FKBP12; PPIASE; FKBP-12; FKBP12C FKBP12.6, FKBP1L, FKBP9, OTK4, PKBP1L, PPIase FKBP51, FKBP54, P54, PPIase, Ptg-10 Bos1, GS27, Membrin

11336 2280

Plasma membrane Broad Cytoplasm, Sarcoplasmic Recticulum Cytoplasm Hypertension

FKBP1B

FK506 binding protein 1B, 12.6 kDa FK506 binding protein 5

2281

Isomerase activity, Muscle contraction, Protein folding FK506 binding, Protein folding, Isomerase activity, Protein binding Receptor activity, Vesicle mediated transport, Transporter activity, Protein transport DNA binding, Protein folding, Protein binding, Transcription, Transcriptional Activity DNA binding, Protein folding, Protein binding, Transcription, Transcriptional Activity DNA binding, Cell development, Transcriptional Activity, Cell proliferation, Protein folding, Transcription Nucleotide binding, Protein folding

Broadly

FKBP5

2289

Broad

Nucleus

Depression

GOSR2

Golgi SNAP receptor complex member 2

9570

Golgi apparatus, Endoplasmic reticulum, Membrane Broadly Cytoplasm, Nucleus Cytoplasm, Nucleus Nucleus

HSF1

Heat Shock transcription factor 1 Heat Shock transcription factor 2 Heat Shock transcription factor 4 Heat Shock 70kDa Protein 4-like Hsp70-interacting Protein KDEL (Lys-Asp-Glu-Leu) endoplasmic reticulum protein retention receptor 1 [Homo sapiens] KDEL (Lys-Asp-Glu-Leu) endoplasmic reticulum protein retention receptor 2 [Homo s apiens] KDEL (Lys-Asp-Glu-Leu) endoplasmic reticulum protein retention receptor 3 [Homo sapiens] Protein disulfide isomerase family A, member 2

HSTF1

3297

HSF2

3298

Broadly

HSF4

CTM

3299

Broad

HSPA4L HSPBP1 KDELR1

APG-1, Osp94

22824 23640

Broad Broad

Cytoplasm, Nucleus

ERD2, ERD2.1, HDEL, PM23

10945

ER retention, Protein retention, Protein transport

Endoplasmic reticulum Golgi apparatus Endoplasmic reticulum Golgi apparatus Endoplasmic reticulum Membrane Endoplasmic reticulum

KDELR2

ELP-1; ERD2.2

11014


KDELR3

ERD2L3

11015


PDIA2

PDA2, PDI, PDIP, PDIR

64714

Isomerase activity, Apoptosis, Protein binding, Redox homeostasis, Protein folding, Protein retention


17

Chaperones & Others (continued)Official Symbol PDIA3 Name Protein disulfide isomerase family A, member 3 Synonyms ERp57, ERp60, ERp61, GRP57, GRP58, P58, PI-PLC Entrez Gene ID 2923 Biological Function Cysteine endopeptidase activity, Redox homeostasis, Phospholipase C activity, Regulation of apoptosis, Protein binding, Protein import into nucleus, Disulfide isomerase activity, Protein retention in ER, Signal transuction Tissue Distribution Broad Cellular Distribution Endoplasmic reticulum Human Diseases

PDIA3P

Protein disulfide isomerase family A, member 3 pseudogene Protein disulfide isomerase family A, member 4 Protein disulfide isomerase family A, member 5 Protein disulfide isomerase family A, member 6 Prostaglandin E synthase 3 (cytosolic)

Erp60, GRP58P

171423

PDIA4

Erp70, Erp72

12304

Ion binding, Redox homeostasis, Isomerase activity, Protein secretion, Protein disulfide isomerase activity Isomerase activity, Electron transport, Oxidoreductase activity, Redox homeostasis, Protein folding Isomerase activity, Redox homeostasis, Protein folding Isomerase activity, Fatty acid biosynthesis, Prostaglandin E sytnhase, Prostanoid biosynthesis, Telomerase activity, Signal transduction, Telomere maintenance Binding, Response to stress, Protein folding Nucleotide binding, Protein folding Endopeptidase activity, Protein folding, Nucleotide binding Nucleotide binding, Protein folding Nucleotide binding, Protein folding Glucosyltransferase activity, Protein folding, Amino acid glysoylation Glucosyltransferase activity, Protein folding, Amino acid glysoylation Broad Broad Broad Broad

Endoplasmic reticulum Endoplasmic reticulum Endoplasmic reticulum Cytoplasm, Nucleus

PDIA5

FLJ30401, PDIR

10954

PDIA6 PTGES3

ERP5, P5, TXNDC7 P23, TEBP

10130 10728

STIP1

Stress-induced-phosphoprotein 1 (Hsp70/Hsp90organizing protein) t-complex 1 torsin family 1, member A (torsin A) torsin family 1, member B (torsin B) TNF receptor-associated Protein 1 UDP-glucose ceramide glucosyltransferase-like 1 UDP-glucose ceramide glucosyltransferase-like 2

HOP; P60; STI1L

10963

Golgi, Nucleus

TCP1 TOR1A

CCT-a, CCT1, CCTa, TCP-1-a DQ2, DYT1, torsin A DQ1 HSP75, HSP90L HUGT1 HUGT2

6950 1861

Cytoplasm Cytoplasmic,, Endoplasmic Reticulum Endoplasmic Reticulum Mitochondrion Endoplasmic reticulum Endoplasmic reticulum Dystonia

TOR1B TRAP1 UGCGL1 UGCGL2

27348 10131 56886 55757

Grp94 Monoclonal Antibody (Cat. No. SPA-850)Human colon cancer CoCa-2 cells were analyzed by flow cytometry using isotype control antibody (left) or Grp94 monoclonal antibody (clone 9G10; right) at a final concentration of 10g/mL.

18

Grp75 Monoclonal Antibody (Cat. No. SPS-825)Human breast cancer tissue was immunohistochemically stained using Grp75 monoclonal antibody (clone 30A5) at a dilution of 1:50.

TCP-1a Monoclonal Antibody (Cat. No. CTA-191)Human colon cancer Coca-2 cells were analyzed by flow cytometry using isotype control antibody (left) or TCP-1a monoclonal antibody (clone 91a; right) at a final concentration of 10g/mL.


19

Hsp27 Reviewexpressed in a variety of tissues; their expression, however, is also up-regulated under conditions of stress as well as in a variety of disease settings14. Members of the sHSP family are categorized on the basis that they possess a conserved C-terminal region known as the a-crystallin domain and a variable N-terminal region. The a-crystallin domain consists of two anti-parallel b-sheets14. It is worth noting that while proteins like HSP32/HO-1 have also been categorized as sHSPs, they lack the critical C-terminal a-crystallin domain defining this family of heat shock proteins. The small heat shock family members vary in their respective molecular weights; they range in size from 15 kDa to 30 kDa. These proteins are known to exist as either homo- or heterocomplexes ranging in size from single units to large multimeric complexes up to approximately 700 kDa3. It is well documented that sHSP family members undergo post-translational modifications with the most common being the phosphorylation of serine residues. For instance, both the human form of aB-crystallin and HSP27 are phosphorylated on three serine residues. In the case of aB-crystallin, this protein is phosphorylated on Ser-19, Ser-45, and Ser-5913; whereas HSP27 is phosphorylated on Ser-15, Ser-78, and Ser-82, respectively16,22. While little homology exists in the sequences flanking these phosphorylation sites, there is definite overlap in regard to the kinases that phosphorylate these sites. In particular, the mitogen activated protein kinase activated protein kinase, MAPKAPK-2 is one of the key protein kinases able to phosphorylate many of these sites with the exception of Ser-19 on aB-crystallin, both in vitro and in vivo. At present, the kinase responsible for phosphorylating Ser-19 of on aB-crystallin has yet to be identified. Other kinases implicated in the phosphorylation of these serine sites are Erk-1 and Erk-2, which phosphorylate aBcrystallin15, and MAPKAPK-3, PKAca, p70S6K, PKD1, and PKCd, which phosphorylate HSP276,9,16,21,22.

HSP27: A regulator of cellular invasion. HSP27 localizes to focal adhesions, influences membrane dynamics and enhances the invasive phenotype of malignant cells.

Heat shock proteins (HSP), also known as molecular chaperones, are critical regulators of cellular homeostasis. Initially identified some forty years ago as heat responsive genes, HSPs have been reported to play important roles in the folding of nascent or new proteins, guiding the renaturation of misfolded or partly denatured proteins, as well as facilitating cellular turnover of client proteins8,12,14. In this regard, HSP90 and its associated co-chaperone complex is known to recruit E3 ubiquitin ligases under certain conditions, favoring the ubiquitinylation and degradation of the client proteins. However, in the presence of ATP, HSP90 complexes can favor the stabilization of these client proteins23. HSPs are categorized into six different families according to their respective molecular weights. They are the HSP100 family, the HSP90 family, the HSP70 family, the HSP60 family, the HSP40 family, and the small heat shock family (sHSPs) including HSP27.

Upstream Signals Leading to the Phosphorylation of HSP27HSP27 has been reported to be phosphorylated in response to a variety of extracellular-derived signals including TNFa, thrombin, bFGF stimulation, as well as under conditions of heat shock and oxidative stress7,16. It is postulated that these pathways converge on and elicit their effects through p38 MAPK and MAPKAPK-2. p38 MAPK has been reported to phosphorylate MAPKAPK-2 in vitro on several residues including Thr-25, Thr-222, and Thr-334 leading to its activation5. Upon activation, MAPKAPK-2 is believed to phosphorylate HSP27 in

Small Heat Shock FamilyThe small heat shock family (sHSP) of molecular chaperones is a ubiquitously expressed group of proteins that is highly conserved among species. To date, ten sHSP isoforms have been identified and designated HSPB1 through HSPB10, respectively, with the most common being HSPB1 or HSP27, and HSPB5 or aB-crystallin14. Both HSP27 and aB-crystallin are constitutively

20

vivo. MAPKAPK-2, as well as MAPKAPK-3, has been reported to phosphorylate HSP27 in vitro on all three HSP27 phosphorylation sites, Ser-15, Ser-78, and Ser-8216,22. Studies utilizing the p38 inhibitor, SB203580, have also offered conclusive evidence that the p38 pathway is intimately involved in the post-translational regulation of HSP27. Treatment with SB203580, is able to attenuate the phosphorylation of HSP27 in response to various agonists18. More recently, both PKD1 and AKT1 have been ascribed as HSP27 phosphorylating kinases. PKD1 was shown to phosphorylate HSP27 in vitro, however, only on Ser-15 and Ser-829. AKT1, on the other hand, was shown to interact with p38 and phosphorylate HSP27 on Ser-8221.

HSP27: Clinical RamificationsThere is a wealth of evidence supporting the notion that heat shock proteins may contribute to the pathogenesis of human diseases like cancer, cardiovascular disease, and neurological disorders. Moreover, elevated expression of certain HSPs, like HSP27, has been reported to correlate with poor patient outcome in breast, ovarian, and prostate cancer8. This might, in part, be due to the effects HSP27 has on the cytoskeleton. Recently, HSP27 has been reported to behave as an actin-capping protein and influence actin dynamics; the latter of which was shown to be dependent upon the phosphorylation status of HSP274,17. In this regard, unphosphorylated monomeric HSP27 was shown to inhibit the polymerization of actin in vitro, whereas multimeric HSP27 complexes appeared to have no inherent effect on actin dynamics regardless of phosphorylation status. HSP27 has also been demonstrated to localize to focal adhesions, influence membrane dynamics, as well as influence invasive phenotype of cells2,11,24. More recently, HSP27 has been shown to play a role in the regulation of matrix metalloproteinase (MMP)-2 activity in prostate cancer cells24. Taken together, it would appear that disrupting the interactions between HSP27 and its binding partners, or reducing its expression in cancer might be a therapeutically valid approach when combined with conventional methodologies.

HSP27: FunctionAs a high molecular weight complex, HSP27 plays a critical role in the renaturation of misfolded or partly denatured proteins by specifically blocking their aggregation10. As with other phospho-proteins, this function is tightly linked with the phosphorylation status of HSP27. Phosphorylation of Ser-82 has been shown to result in HSP27 complex dissociation and the subsequent loss of its chaperoning activity. In addition to its chaperoning function, HSP27 has been shown to interact with different cytoskeletal elements affecting actin polymerization4 as well as inhibiting apoptosis7,19,20. Apoptosis, or programmed cell death, is a finely coordinated process involving the activation of a discrete network of enzymes, referred to as cellular caspases, that aid in preserving the fidelity of the human genome as well as turning over damaged or worn-out cells. While there is variation in terms of the mechanism by which apoptosis can be elicited (e.g., death receptor apoptosis vs. mitochondrial mediated apoptosis), these pathways converge on the key executioners of apoptosis, caspases-3, 6 and 7, to carry out the process. To counteract these pro-apoptotic mechanisms, the cell has devised a number of ways to inhibit this process. Two of the best-defined mechanisms involve the overexpression of the B-cell lymphoma protein, Bcl-2 and the activation of the anti-apoptotic kinase, AKT. More recently, HSP27 has also been ascribed as an anti-apoptotic protein. In particular, HSP27 has been reported to inhibit apoptosis (1) through its interactions with the death associated protein DAXX7, (2) by facilitating the activation of AKT21, and (3) by blocking the formation of the apoptosome19,20. Taken together, the overexpression of HSP27 in the context of a disease such as cancer, would facilitate adaptation to stressful conditions by aiding in the suppression of apoptosis, ultimately leading to a more aggressive phenotype. As such, it is not surprising that the overexpression of HSP27 correlates with poor patient prognosis in a variety of lesions.

HSP27: Phosphorylation linked to function. HSP27 is phosphorylated on key serine residues by MAPKAPK2 as well as MAPKAPK3 amongst others. Phosphorylation is associated with the dimerization of HSP27 and its function.

References on page 26

21

Hsp70 ReviewInitially identified by Ritossa some forty years ago, heat shock proteins (HSP) were first identified as genes up-regulated in response to heat shock stimulation in Drosophila20. They are now recognized as proteins that play critical roles in cellular homeostasis and the adaptation to stressful conditions such as heat shock, oxidative stress, genotoxic shock, viral infection, and hypoxic conditions26. In part, the cytoprotective effects of HSPs are achieved through their role in the re-folding of partly denatured or misfolded proteins. HSPs are also known to be involved in: (1) the folding of newly formed proteins, (2) the trafficking of cellular proteins, as well as (3) the turn over of cellular proteins through the proteasomal pathway; as such, HSPs have been classified as molecular chaperones. HSPs are ubiquitously expressed and highly conserved among species, ranging from the simplest prokaryotes to complex eukaryotes such as humans. They are classified according to their respective molecular weights and are divided into six families: the small HSPs (sHSPs), the HSP40 family, the HSP60 family, the HSP70 family, the HSP90 family, and the HSP100 family. normal cellular function. Two other members of the HSP70 family under active investigation are the endoplasmic reticulum(ER) and mitochondrial-associated members, referred to as the glucose regulated proteins, GRP78 and GRP75. GRP78 plays a critical role in the ER-associated stress response, whereas GRP75 (also known as mortalin) is involved in the maintenance of mitochondrial function.

HSP70: Structure and FunctionAll HSP70 family members contain a highly conserved N-terminal ATPase domain of approximately 44 kDa which possesses weak ATPase activity under normal conditions. These proteins also contain an approximately 25 kDa C-terminal region which consists of a conserved hydrophobic peptide binding domain (PBD) of approximately 15 kDa, and a more variable a-helical domain of approximately 10 kDa5. The a-helical domain is classified as a cap believed to open and close in response to changes in the nucleotide binding status of HSP7015. The activity and function of HSP70 are thought to result from the binding of ATP to the N-terminus. In its ATP-bound state, the C-terminus of HSP70 is said to undergo a conformational change, resulting in the opening of the a-helical cap; this opening allows substrates to interact with the hydrophobic pocket of the PBD. In its ADP-bound state, it is believed that the a-helical cap is closed, excluding substrates from the PBD. Thus, in the presence of ATP, HSP70 is said to favor the folding of its client protein, as it has a lower affinity and faster processing rate than the ADP-bound form5,15. However, unlike other enzyme classes, members of the HSP70 family bind to a wide variety of client proteins through short hydrophobic segments found in unfolded, misfolded, or denatured proteins. On its own, members of the HSP70 family are known to possess little or no intrinsic ATPase activity. As with other heat shock proteins, ATPase activity and function are thought to be influenced through interactions with specific co-chaperone molecules. HSP70 co-chaperones include: (1) HSP40, which is believed to assist in loading targets on the HSP70 machinery3, (2) the HSC70 interacting protein, Hip, which binds to the ATPase domain stimulating its activity3, (3) the HSC70/HSP90 organizing protein, Hop, which serves as a link between HSP70 and HSP90 allowing for substrate exchange between the two chaperones, and also facilitates ATP/ADP cycling3, (4) the HSC70 unbinding protein, Hup, which facilitates the release of unfolded proteins3 , (5) the HSC70 accessory protein, Hap3, and the Bcl-2 associated athanogene proteins, BAG-1, BAG-2, and BAG-3, which interact with the ATPase domain and block the binding of unfolded proteins4,11,23,27, and (6) the carboxyl-terminus of HSP70 interacting protein, CHIP, which is believed to be a bona

The HSP70 FamilyThe HSP70 family represents one of the most widely examined heat shock families and consists of up to ten highly homologous members. Refer to Table 1 for a representative list of the HSP70 family members. As with other heat shock families, members of the HSP70 family differ in their spatial and subcellular distribution, as well as their expression levels under normal, unstressed conditions21,24. It is well established that the expression of some members of the HSP70 family can be induced under conditions of stress; as these proteins do not contain introns, they are able to be quickly up-regulated. While the expression of many HSP70 family members can be up-regulated in response to various stressors, the major stress inducible HSP70 members are the highly homologous genes HSP1A1 and HSPA1B, also referred to as HSP70-1 and HSP70-2, respectively. These proteins are expressed at relatively low or undetectable levels in most normal, unstressed cells; however, upon insult, their expression dramatically increases. It is worth noting, that HSPA6 and HSPA7, the two other highly related HSP70 inducible members known as HSP70B and HSP70B are up-regulated only under conditions of extreme stress21,24. At present, further research is required to elucidate the physiological importance of these two HSP70 family members. The constitutively expressed member of HSP70 family is HSPA8, also referred to as HSC70. HSC70 is ubiquitously expressed in all cell types and is believed to be responsible for maintaining

22

fide E3 ubiquitin ligase assisting in the ubiquitinylation of cellular proteins1,10,16. Overall, members of the HSP70 family are known to play critical roles in the folding of newly synthesized proteins; the re-folding of misfolded or denatured proteins; the trafficking of proteins across cellular membranes; the disassembly of clathrin-coated vesicles; the inhibition of protein aggregation; and the targeting and degradation of proteins via the proteasomal pathway3. More recently, HSP70 has also been demonstrated to suppress apoptosis in response to various stimuli12.

HSP70: Effects on ApoptosisApoptosis is a highly coordinated process that functions either through the activation of a discrete network of cysteine proteases known as cellular caspases, or through mechanisms not depending on caspase activation, known as caspase-independent cell death. The activation of cellular caspases can be achieved through various mechanisms including death signals provided at the level of the cell membrane, the mitochondria, and the endoplasmic reticulum. While these pathways rely on different initiators of apoptosis, they all converge on, and elicit their effects through, the executioner caspases mainly caspase-3, -6 and -7. Along with the activation of endonucleases, the activation of these caspases results in the dismantling of the cell and its intracellular contents, leading to the eventual formation of apoptotic bodies.

It is well established that the commitment to undergo apoptosis can be attenuated or blocked through various mechanisms, including the activation of key signal transduction molecules such as Akt, and the overexpression of certain cellular proteins such as Bcl-2. More recently, overexpression of heat shock proteins mainly members of the HSP70 family has also been demonstrated to inhibit apoptosis. Elevated expression of HSP70 has been reported to: (1) inhibit the formation of the apoptosome2, (2) inhibit the translocation of the Bcl-2 protein, Bax, to the mitochondrial membrane, ultimately blocking the release of cytochrome c from the mitochondria22, (3) inhibit the activation of cellular caspases19, (4) block the activation of the apoptosis signal regulating kinase, ASK118, (5) inhibit the activation of the stress kinase p388, and (6) inhibit JNK activation17.

HSP70: Clinical Ramifications in CancerThere is a wealth of literature supporting the notion that heat shock proteins are elevated in a variety of human malignancies and that these increases in expression may not only contribute to the pathogenesis of the disease25, but may be of diagnostic, prognostic and therapeutic importance6. In this regard, HSP70 has been reported to correlate with poor differentiation in certain malignancies, increased lymph node metastases, chemo-resistance, and poor clinical outcome6,14. Further studies evaluating the importance of HSP70 family members in the initiation and progression of cancer will undoubtedly be of great clinical importance.

HSP70: A cell survival protein. HSP70 suppresses apoptosis by inhibiting the formation of the apoptosome and by blocking the activation of stress induced kinases including: ASK1, p38, and JNK, respectively.

References on page 26


23

Hsp90 Reviewleft: HSP90: A regulator of cell survival. Inhibition of HSP90 activity by drugs like geldanmycin destabilize client proteins which ultimately leads to the onset of apoptosis. right: HSP90: A drugable target. Inhibition of HSP90 by geldanmycin (GA) favors the ubiquitinylation and degradation of client proteins.

The heat shock, or stress family of proteins is a highly conserved, ubiquitously expressed class of proteins that have been demonstrated to be intimately involved in the regulation of cellular homeostasis in response to a myriad of environmental and physiological stressors27. The heat shock proteins (HSP) are classified into six different groups according to their respective molecular weights; they are the small heat shock proteins including HSP27, the HSP40 family, the HSP60 family, the HSP70 family, the HSP90 family, and the HSP100 family, respectively. HSPs, commonly referred to as molecular chaperones, were initially identified and described over thirty years ago as heat shock responsive genes. HSPs are now know to play critical roles in the stabilization of partly denatured or misfolded proteins, facilitate proper folding of nascent or new polypeptides as well as regulate the spatial distribution of cellular proteins. More recently, these molecular chaperones, mainly HSP90, have received significant attention for the putative roles they play in the pathogenesis and progression of human diseases like cancer21,35.

All members of the HSP90 family possess three specific domains: an N-terminal nucleotide binding pocket to which most clinical compounds are being developed6,22, a central domain important for ATPase activity11,19, and a C-terminal domain believed to act as a second nucleotide binding site9. HSP90 can either exist as a homodimer, a heterodimer, or as a multiprotein complex with other co-chaperones including HSP40, HSP70, Hop, and p2328. As with other heat shock families, dimerization is believed to be an ATP-dependent process. Unlike other member; however, the N-terminal nucleotide binding site of HSP90 is highly unique and bears a strong resemblance to members of the GKHL superfamily including bacterial gyrase, MutL and histidine kinases8. The mechanism(s) involved in regulating HSP90 activity and function, is at present unclear. However thought to involve: (1) post-translational modification including acetylation and phosphorylation1,17,20, (2) the N-terminal nucleotide binding status28, and (3) interactions with accessory co-chaperones28. Early studies evaluating the phosphorylation status of HSP90 revealed that phosphorylation of this target is essential for its activity. In this regard, HSP90 has been reported to be tyrosine phosphorylated in vivo when complexed with other proteins1. These specific phosphorylation sites have yet to be elucidated, however. More recently, the acetylation status of HSP90 has also been reported to influence the activity of HSP90. In particular, these studies revealed that hyperacetylation of HSP90 led to decrease in its association with the essential co-chaperone, p23, and a concomitant loss of chaperoning activity20. The specific acetylation sites have been yet to be reported. The N-terminal nucleotide binding status has also been demonstrated to influence the function of HSP90. In the presence of ATP and the appropriate upstream signals, HSP90 cyclizes with its co-chaperone molecules, stabilizing the expression of its client proteins. However, when bound by inhibitors like

HSP90: Structure and FunctionThe HSP90 family, which consists of both inducible and constitutive isoforms, is encoded at two distinct loci. Cellular homologues of the HSP90 family include: HSP90a (the inducible isoform), HSP90b (the constitutive isoform) Grp94 and Trap1, as well as the recently identified variant HSP90N. Whereas, HSP90a and HSP90b are predominantly cytosolic proteins and represent 1-2% of the normal cellular protein content, Grp94 and Trap1 are localized within the endoplasmic reticulum (ER) and mitochondria, respectively, and are expressed at much lower levels than their cytoplasmic counterparts28. HSP90N, on the other hand, preferentially localizes to the cellular membrane through its unique N-terminal hydrophobic region10.

24

Geldanamycin (GA), HSP90 function is impaired which results in the recruitment of E3 ubiquitin ligases favoring the ubiquitinylation and degradation of client proteins by the 26S proteasomal complex28. The stabilization versus degradation of proteins, in the presence of physiological stressors, may tip the balance in favor of stabilizing mutant proteins ultimately promoting an anti-apoptotic or pro-survival state. As with other heat shock proteins, HSP90 requires a number of co-chaperone molecules for its full function including HSP70, HSP40, Hip/Hop, p23, immunophilins, and CDC37/p5028. More recently, Aha1 (Activator of HSP90 ATPase homologue 1), which associates with the central domain of HSP90, was identified as a key molecular co-chaperone required for ATPase function18.

p53 has been marked as the master regulator of the human genome, knocking out this function of this essential protein through stabilization of the mutated form allows for the propagation of additional favorable oncogenic mutations, including those that facilitate progression of the disease e.g., invasion and metastasis.

HSP90: Clinical Ramifications in CancerSeveral lines of evidence support the notion that the expression levels of heat shock proteins are dramatically increased in a variety of human malignancies, both in solid and hematological types, and might contribute to the advancement of the disease3,5,7,12,15,16,21,23,24,26,34,35. As an example, elevated levels of HSP27 have been reported to correlate with poor patient outcome in prostate, breast, and ovarian cancer4. Moreover, compelling evidence suggests that HSP90, in the context of cancer, exists in a hyperactive state in the diseased tissue when compared with the normal surrounding material14. In its highly active state, the HSP90 complex would theoretically influence key signaling nexuses that would contribute to the pathogenesis of the disease; these include cell growth promotion, evasion of apoptosis, and stimulation of angiogenesis. In this regard, it has been postulated that changes in expression and activity levels are critical in that they provide a mechanism to counteract, or compensate, for the selective pressures these abnormal cells experience as the disease progresses including gene mutations, hypoxia, and growth pressures amongst others. As such, heat shock proteins, mainly HSP90, appear to provide a buffering mechanism facilitating the natural selection of the strongest cell lineage ultimately allowing for adaptation to the harshest of conditions. Consequently, based upon the uniqueness of the N-terminal nucleotide binding site, the differences in biological activity between the cancer and normal tissue, and the notion that HSP90 stabilizes key oncogenic client proteins, HSP90 has been suggested to serve as an excellent target for therapeutic intervention2,13,31. Not only is targeting HSP90 selecting against those adaptable cell lineages, but in addition, key oncogenic proteins would be destabilized and consequently degraded, shifting the balance towards an anti-proliferative pro-apoptotic state. Currently, there are on-going clinical trials evaluating the efficacy of these N-terminal specific inhibitors and a number of clinical candidates in the pipeline. Perhaps, the future will entail a combinatorial modality involving the administration of selective HSP90 inhibitors with conventional methodologies. References on page 26

HSP90: Client ProteinsIn comparison to other members of the heat shock family, HSP90 client proteins are unique and tend to encompass key signal transduction molecules involved in the regulation of cellular growth, survival, and differentiation. In this regard, one of the first client proteins to be described was the transforming tyrosine kinase isolated from the Rous sarcoma virus, v-Src30,33. Since this original discovery, the number of client proteins influenced by HSP90 has significantly increased, many of which are linked with the pathogenesis of human cancer. Client proteins include serine/threonine and tyrosine kinases, transcription factors, and steroid receptors as well as certain tumor suppressor proteins. A concise list of clients influenced by HSP90 include: Akt/PKB, ASK1, Aurora B, Bcr-Abl, CDK1, CDK4, CHK1, CKII, ErbB2/Her-2, EGFR, ER, Hif-1a, c-Met, mutant p53, PLK, and Raf28. Two key signaling molecules influenced by HSP90, which are central to normal physiology, are the protein kinase Akt, and the tumor suppressor protein p53. In the case of Akt, HSP90 has been demonstrated to bind with the phosphorylated form and protect it from being inactivated by its dephosphorylating phosphatase, protein phosphatase (PP)-2A25. When active, Akt is known to provide anti-apoptotic signals, through various mechanisms, ultimately preventing the cells from undergoing apoptosis. In this regard, Akt in association with HSP90, has been demonstrated to phosphorylate and inhibit the apoptotic kinase ASK1, thus blocking apoptosis36. In the case of p53, HSP90 has been reported to bind with and stabilize the expression of the mutated form of this protein29. Stabilization of the mutated form provides a mechanism of knocking out the function of the normal wild-type p53 molecules through a dominant-negative effect. Functional p53 exists as a tetramer; as such, one mutated copy of p53 within this complex is capable of impairing normal p53 function. As

25

ReferencesHsp27 Referencesheat shock proteins. FEBS Lett. 1992 Nov 30;313(3):307-313. 23. Whitesell L, Lindquist SL. HSP90 and the chaperoning of cancer. Nat Rev Cancer. 2005 Oct;5(10):761-772. 24. Xu L, Chen S, Bergan RC. MAPKAPK2 and HSP27 are downstream effectors of p38 MAP kinase-mediated matrix metalloproteinase type 2 activation and cell invasion in human prostate cancer. Oncogene. 2006 May 18;25(21):2987-2998. 1. Aldrian S, Trautinger F, Frohlich I, Berger W, Micksche M, Kindas-Mugge I. Overexpression of Hsp27 affects the metastatic phenotype of human melanoma cells in vitro. Cell Stress Chaperones. 2002 Apr;7(2):177-185. 2. Aldrian S, Kindas-Mugge I, Trautinger F, Frohlich I, Gsur A, Herbacek I, Berger W, Micksche M. Overexpression of Hsp27 in a human melanoma cell line: regulation of Ecadherin, MUC18/MCAM, and plasminogen activator (PA) system. Cell Stress Chaperones. 2003 Fall;8(3):249-57. 3. Beck FX, Neuhofer W, Muller E. Molecular chaperones in the kidney: distribution, putative roles, and regulation. Am J Physiol Renal Physiol. 2000 Aug;279(2):F203-F215. 4. Benndorf R, Hayess K, Ryazantsev S, Wieske M, Behlke J, Lutsch G. Phosphorylation and supramolecular organization of murine small heat shock protein HSP25 abolish its actin polymerization-inhibiting activity. J Biol Chem. 1994 Aug 12;269(32):20780-20784. 5. Ben-Levy R, Leighton IA, Doza YN, Attwood P, Morrice N, Marshall CJ, Cohen P. Identification of novel phosphorylation sites required for activation of MAPKAP kinase-2. EMBO J. 1995 Dec 1;14(23):5920-5930. 6. Butt E, Immler D, Meyer HE, Kotlyarov A, Laass K, Gaestel M. Heat shock protein 27 is a substrate of cGMP-dependent protein kinase in intact human platelets: phosphorylation-induced actin polymerization caused by HSP27 mutants. J Biol Chem. 2001 Mar 9;276(10):7108-7113. 7. Charette SJ, Lavoie JN, Lambert H, Landry J. Inhibition of Daxx-mediated apoptosis by heat shock protein 27. Mol Cell Biol. 2000 Oct;20(20):7602-7612. 8. Ciocca DR, Calderwood SK. Heat shock proteins in cancer: diagnostic, prognostic, predictive, and treatment implications. Cell Stress Chaperones. 2005 Summer;10(2):86 103. 9. Doppler H, Storz P, Li J, Comb MJ, Toker A. A phosphorylation state-specific antibody recognizes Hsp27, a novel substrate of protein kinase D. J Biol Chem. 2005 Apr 15;280(15):15013-15019. 10. Ehrnsperger M, Graber S, Gaestel M, Buchner J. Binding of non-native protein to Hsp25 during heat shock creates a reservoir of folding intermediates for reactivation. EMBO J. 1997 Jan 15;16(2):221-229. 11. El-Ghobashy AA, Shaaban AM, Innes J, Prime W, Herrington CS. Upregulation of heat shock protein 27 in metaplastic and neoplastic lesions of the endocervix. Int J Gynecol Cancer. 2005 May-Jun;15(3):503-509. 12. Haslbeck M, Franzmann T, Weinfurtner D, Buchner J. Some like it hot: the structure and function of small heat-shock proteins. Nat Struct Mol Biol. 2005 Oct;12(10):842-846. 13. Ito H, Okamoto K, Nakayama H, Isobe T, Kato K. Phosphorylation of aB-crystallin in response to various types of stress. J Biol Chem. 1997 Nov 21;272(47):29934-29941. 14. Kappe G, Franck E, Verschuure P, Boelens WC, Leunissen JA, de Jong WW. The human genome encodes 10 a-crystallin-related small heat shock proteins: HspB1-10. Cell Stress Chaperones. 2003 Spring;8(1):53-61. 15. Kato K, Ito H, Kamei K, Inaguma Y, Iwamoto I, Saga S. Phosphorylation of aB-crystallin in mitotic cells and identification of enzymatic activities responsible for phosphorylation. J Biol Chem. 1998 Oct 23;273(43):28346-28354. 16. Landry J, Lambert H, Zhou M, Lavoie JN, Hickey E, Weber LA, Anderson CW. Human HSP27 is phosphorylated at serines 78 and 82 by heat shock and mitogen-activated kinases that recognize the same amino acid motif as S6 kinase II. J Biol Chem. 1992 Jan 15;267(2):794-803. 17. Miron T, Vancompernolle K, Vandekerckhove J, Wilchek M, Geiger B. A 25-kD inhibitor of actin polymerization is a low molecular mass heat shock protein. J Cell Biol. 1991 Jul;114(2):255-261. 18. Muller E, Burger-Kentischer A, Neuhofer W, Fraek ML, Marz J, Thurau K, Beck FX. Possible involvement of heat shock protein 25 in the angiotensin II-induced glomerular mesangial cell contraction via p38 MAP kinase. J Cell Physiol. 1999 Dec;181(3):462-469. 19. Pandey P, Farber R, Nakazawa A, Kumar S, Bharti A, Nalin C, Weichselbaum R, Kufe D, Kharbanda S. Hsp27 functions as a negative regulator of cytochrome c-dependent activation of procaspase-3. Oncogene. 2000 Apr 13;19(16):1975-1981. 20. Paul C, Manero F, Gonin S, Kretz-Remy C, Virot S, Arrigo AP. Hsp27 as negative regulator of cytochrome C release. Mol Cell Biol. 2002 Feb;22(3):816-834. 21. Rane MJ, Pan Y, Singh S, Powell DW, Wu R, Cummins T, Chen Q, McLeish KR, Klein JB. Heat shock protein 27 controls apoptosis by regulating Akt activation. J Biol Chem. 2003 Jul 25;278(30):27828-27835. 22. Stokoe D, Engel K, Campbell DG, Cohen P, Gaestel M. Identification of MAPKAP kinase 2 as a major enzyme responsible for the phosphorylation of the small mammalian

Hsp70 References

1. Ballinger CA, Connell P, Wu Y, Hu Z, Thompson LJ, Yin LY, Patterson C. Identification of CHIP, a novel tetratricopeptide repeat-containing protein that interacts with heat shock proteins and negatively regulates chaperone functions. Mol Cell Biol. 1999 Jun;19(6):4535-4545. 2. Beere HM, Wolf BB, Cain K, Mosser DD, Mahboubi A, Kuwana T, Tailor P, Morimoto RI, Cohen GM, Green DR. Heat-shock protein 70 inhibits apoptosis by preventing recruitment of procaspase-9 to the Apaf-1 apoptosome. Nat Cell Biol. 2000 Aug;2(8):469-475. 3. Benjamin IJ, McMillan DR. Stress (heat shock) proteins: molecular chaperones in cardiovascular biology and disease. Circ Res. 1998 Jul 27;83(2):117-132. 4. Bimston D, Song J, Winchester D, Takayama S, Reed JC, Morimoto RI. BAG-1, a negative regulator of Hsp70 chaperone activity, uncouples nucleotide hydrolysis from substrate release. EMBO J. 1998 Dec 1;17(23):6871-6878. 5. Bukau B, Horwich AL. The Hsp70 and Hsp60 chaperone machines. Cell. 1998 Feb 6;92(3):351-366. 6. Ciocca DR, Calderwood SK. Heat shock proteins in cancer: diagnostic, prognostic, predictive, and treatment implications. Cell Stress Chaperones. 2005 Jun; 10(2): 86-103. 7. Freeman BC, Myers MP, Schumacher R, Morimoto RI. Identification of a regulatory motif in Hsp70 that affects ATPase activity, substrate binding and interaction with HDJ-1. EMBO J. 1995 May 15;14(10):2281-2292. 8. Gabai VL, Meriin AB, Mosser DD, Caron AW, Rits S, Shifrin VI, Sherman MY. Hsp70 prevents activation of stress kinases. A novel pathway of cellular thermotolerance. J Biol Chem. 1997 Jul 18;272(29):18033-18037. 9. Hohfeld J, Minami Y, Hartl FU. Hip, a novel cochaperone involved in the eukaryotic Hsc70/Hsp40 reaction cycle. Cell. 1995 Nov 17;83(4):589-98. 10. Kampinga HH, Kanon B, Salomons FA, Kabakov AE, Patterson C. Overexpression of the cochaperone CHIP enhances Hsp70-dependent folding activity in mammalian cells. Mol Cell Biol. 2003 Jul;23(14):4948-4958. 11. Kanelakis KC, Morishima Y, Dittmar KD, Galigniana MD, Takayama S, Reed JC, Pratt WB. Differential effects of the hsp70-binding protein BAG-1 on glucocorticoid receptor folding by the hsp90-based chaperone machinery. J Biol Chem. 1999 Nov 26;274(48):34134-34140. 12. Klein SD, Brune B. Heat-shock protein 70 attenuates nitric oxide-induced apoptosis in RAW macrophages by preventing cytochrome c release. Biochem J. 2002 Mar 15;362(Pt 3):635-641. 13. Li CY, Lee JS, Ko YG, Kim JI, Seo JS. Heat shock protein 70 inhibits apoptosis downstream of cytochrome c release and upstream of caspase-3 activation. J Biol Chem. 2000 Aug 18;275(33):25665-25671. 14. Luk JM, Lam CT, Siu AF, Lam BY, Ng IO, Hu MY, Che CM, Fan ST. Proteomic profiling of hepatocellular carcinoma in Chinese cohort reveals heat-shock proteins (Hsp27, Hsp70, GRP78) up-regulation and their associated prognostic values. Proteomics. 2006 Feb;6(3):1049-1057. 15. Moro F, Fernandez-Saiz V, Slutsky O, Azem A, Muga A. Conformational properties of bacterial DnaK and yeast mitochondrial Hsp70. Role of the divergent C-terminal a-helical subdomain. FEBS J. 2005 Jun;272(12):3184-3196. 16. Murata S, Minami Y, Minami M, Chiba T, Tanaka K. CHIP is a chaperone-dependent E3 ligase that ubiquitylates unfolded protein. EMBO Rep. 2001 Dec;2(12):1133-1138. 17. Park HS, Cho SG, Kim CK, Hwang HS, Noh KT, Kim MS, Huh SH, Kim MJ, Ryoo K, Kim EK, Kang WJ, Lee JS, Seo JS, Ko YG, Kim S, Choi EJ. Heat shock protein hsp72 is a negative regulator of apoptosis signal-regulating kinase 1. Mol Cell Biol. 2002 Nov;22(22):77217730. 18. Park HS, Lee JS, Huh SH, Seo JS, Choi EJ. Hsp72 functions as a natural inhibitory protein of c-Jun N-terminal kinase. EMBO J. 2001 Feb 1;20(3):446-456. 19. Reddy RK, Mao C, Baumeister P, Austin RC, Kaufman RJ, Lee AS. Endoplasmic reticulum chaperone protein GRP78 protects cells from apoptosis induced by topoisomerase inhibitors: role of ATP binding site in suppression of caspase-7 activation. J Biol Chem.

26

2003 Jun 6;278(23):20915-20924. 20. Ritossa F. A new puffing pattern induced by temperature shock and DNP in drosophila. Experentia. 1962 18(12):571-573. 21. Rohde M, Daugaard M, Jensen MH, Helin K, Nylandsted J, Jaattela M. Members of the heat-shock protein 70 family promote cancer cell growth by distinct mechanisms. Genes Dev. 2005 Mar 1;19(5):570-582. 22. Stankiewicz AR, Lachapelle G, Foo CP, Radicioni SM, Mosser DD. Hsp70 inhibits heat-induced apoptosis upstream of mitochondria by preventing Bax translocation. J Biol Chem. 2005 Nov 18;280(46):38729-38739. 23. Takayama S, Bimston DN, Matsuzawa S, Freeman BC, Aime-Sempe C, Xie Z, Morimoto RI, Reed JC. BAG-1 modulates the chaperone activity of Hsp70/Hsc70. EMBO J. 1997 Aug 15;16(16):4887-4896. 24. Tavaria M, Gabriele T, Kola I, Anderson RL. A hitchhikers guide to the human Hsp70 family. Cell Stress Chaperones. 1996 Apr;1(1):23-28. 25. Wadhwa R, Takano S, Kaur K, Deocaris CC, Pereira-Smith OM, Reddel RR, Kaul SC. Upregulation of mortalin/mthsp70/Grp75 contributes to human carcinogenesis. Int J Cancer. 2006 Jun 15;118(12):2973-2980. 26. Wegele H, Muller L, Buchner J. Hsp70 and Hsp90--a relay team for protein folding. Rev Physiol Biochem Pharmacol. 2004;151:1-44. 27. Zeiner M, Gebauer M, Gehring U. Mammalian protein RAP46: an interaction partner and modulator of 70 kDa heat shock proteins. EMBO J. 1997 Sep 15;16(18):5483-5490.

15. Kaur J, Ralhan R. Differential expression of 70-kDa heat shock-protein in human oral tumorigenesis. Int J Cancer. 1995 Dec 11;63(6):774-9. 16. Kimura E, Enns RE, Alcaraz JE, Arboleda J, Slamon DJ, Howell SB. Correlation of the survival of ovarian cancer patients with mRNA expression of the 60-kD heat-shock protein HSP-60. J Clin Oncol. 1993 May;11(5):891-8. 17. Lees-Miller SP, Anderson CW. Two human 90-kDa heat shock proteins are phosphorylated in vivo at conserved serines that are phosphorylated in vitro by casein kinase II. J Biol Chem 1989 Feb; 264(5): 2431-7 18. Meyer P, Prodromou C, Liao C, Hu B, Mark Roe S, Vaughan CK, Vlasic I, Panaretou B, Piper PW, Pearl LH. Structural basis for recruitment of the ATPase activator Aha1 to the Hsp90 chaperone machinery. EMBO J. 2004 Feb 11;23(3):511-9. 19. Meyer P, Prodromou C, Hu B, Vaughan C, Roe SM, Panaretou B, Piper PW, Pearl LH. Structural and functional analysis of the middle segment of hsp90: implications for ATP hydrolysis and client protein and cochaperone interactions. Mol Cell. 2003 Mar;11(3):64758. 20. Murphy PJ, Morishima Y, Kovacs JJ, Yao TP, Pratt WB. Regulation of the dynamics of hsp90 action on the glucocorticoid receptor by acetylation/deacetylation of the chaperone. J Biol Chem. 2005 Oct 7;280(40):33792-9. 21. Nanbu K, Konishi I, Mandai M, Kuroda H, Hamid AA, Komatsu T, Mori T. Prognostic significance of heat shock proteins HSP70 and HSP90 in endometrial carcinomas. Cancer Detect Prev. 1998;22(6):549-55. 22. Prodromou C, Roe SM, OBrien R, Ladbury JE, Piper PW, Pearl LH. Identification and structural characterization of the ATP/ADP-binding site in the Hsp90 molecular chaperone. Cell. 1997 Jul 11;90(1):65-75. 23. Ralhan R, Kaur J. Differential expression of Mr 70,000 heat shock protein in normal, premalignant, and malignant human uterine cervix. Clin Cancer Res. 1995 Oct;1(10):1217-22. 24. Santarosa M, Favaro D, Quaia M, Galligioni E. Expression of heat shock protein 72 in renal cell carcinoma: possible role and prognostic implications in cancer patients. Eur J Cancer. 1997 May;33(6):873-7. 25. Sato S, Fujita N, Tsuruo T. Modulation of Akt kinase activity by binding to Hsp90. Proc Natl Acad Sci U S A. 2000 Sep 26;97(20):10832-7. 26. Trieb K, Gerth R, Holzer G, Grohs JG, Berger P, Kotz R. Antibodies to heat shock protein 90 in osteosarcoma patients correlate with response to neoadjuvant chemotherapy. Br J Cancer. 2000 Jan;82(1):85-7. 27. Wegele H, Muller L, Buchner J. Hsp70 and Hsp90--a relay team for protein folding. Rev Physiol Biochem Pharmacol. 2004;151:1-44. 28. Whitesell L, Lindquist SL. HSP90 and the chaperoning of cancer. Nat Rev Cancer. 2005 Oct;5(10):761-72. 29. Whitesell L, Sutphin PD, Pulcini EJ, Martinez JD, Cook PH. The physical association of multiple molecular chaperone proteins with mutant p53 is altered by geldanamycin, an hsp90-binding agent. Mol Cell Biol. 1998 Mar;18(3):1517-24. 30. Whitesell L, Mimnaugh EG, De Costa B, Myers CE, Neckers LM. Inhibition of heat shock protein HSP90-pp60v-src heteroprotein complex formation by benzoquinone ansamycins: essential role for stress proteins in oncogenic transformation. Proc Natl Acad Sci U S A. 1994 Aug 30; 91(18): 8324-8328. 31. Workman P. Altered states: selectively drugging the Hsp90 cancer chaperone. Trends Mol Med. 2004 Feb;10(2):47-51. 32. Xu W, Marcu M, Yuan X, Mimnaugh E, Patterson C, Neckers L. Chaperone-dependent E3 ubiquitin ligase CHIP mediates a degradative pathway for c-ErbB2/Neu. Proc Natl Acad Sci U S A. 2002 Oct 1;99(20):12847-52. 33. Xu Y, Singer MA, Lindquist S. Maturation of the tyrosine kinase c-src as a kinase and as a substrate depends on the molecular chaperone Hsp90. Proc Natl Acad Sci U S A. 1999 Jan 5;96(1):109-14. 34. Yano M, Naito Z, Tanaka S, Asano G. Expression and roles of heat shock proteins in human breast cancer. Jpn J Cancer Res. 1996 Sep;87(9):908-15. 35. Yufu Y, Nishimura J, Nawata H. High constitutive expression of heat shock protein 90 a in human acute leukemia cells. Leuk Res. 1992 Jun-Jul;16(6-7):597-605. 36. Zhang R, Luo D, Miao R, Bai L, Ge Q, Sessa WC, Min W. Hsp90-Akt phosphorylates ASK1 and inhibits ASK1-mediated apoptosis. Oncogene. 2005 Jun 2;24(24):3954-63.

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1. Adinolfi E, Kim M, Young MT, Di Virgilio F, Surprenant A. Tyrosine phosphorylation of HSP90 within the P2X7 receptor complex negatively regulates P2X7 receptors. J Biol Chem. 2003 Sep 26;278(39):37344-51. 2. Bagatell R, Whitesell L. Altered Hsp90 function in cancer: a unique therapeutic opportunity. Mol Cancer Ther. 2004 Aug;3(8):1021-30. 3. Chant ID, Rose PE, Morris AG. Analysis of heat-shock protein expression in myeloid leukaemia cells by flow cytometry. Br J Haematol. 1995 May;90(1):163-8. 4. Ciocca DR, Calderwood SK. Heat shock proteins in cancer: diagnostic, prognostic, predictive, and treatment implications. Cell Stress Chaperones. 2005 Jun; 10(2): 86-103. 5. Ciocca DR, Clark GM, Tandon AK, Fuqua SA, Welch WJ, McGuire WL. Heat shock protein hsp70 in patients with axillary lymph node-negative breast cancer: prognostic implications. J Natl Cancer Inst. 1993 Apr 7;85(7):570-4. 6. Chadli A, Bouhouche I, Sullivan W, Stensgard B, McMahon N, Catelli MG, Toft DO. Dimerization and N-terminal domain proximity underlie the function of the molecular chaperone heat shock protein 90. Proc Natl Acad Sci U S A. 2000 Nov 7; 97(23): 1252412529. 7. Conroy SE, Sasieni PD, Fentiman I, Latchman DS. Autoantibodies to the 90kDa heat shock protein and poor survival in breast cancer patients. Eur J Cancer. 1998 May;34(6):942-3. 8. Dutta R, Inouye M. GHKL, an emergent ATPase/kinase superfamily. Trends Biochem Sci. 2000 Jan;25(1):24-8. 9. Garnier C, Lafitte D, Tsvetkov PO, Barbier P, Leclerc-Devin J, Millot JM, Briand C, Makarov AA, Catelli MG, Peyrot V. Binding of ATP to heat shock protein 90: evidence for an ATP-binding site in the C-terminal domain. J Biol Chem. 2002 Apr 5;277(14):12208-14. 10. Grammatikakis N, Vultur A, Ramana CV, Siganou A, Schweinfest CW, Watson DK, Raptis L. The role of Hsp90N, a new member of the

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