ActivityScheduled Action/ timing Teaching Assistant Introduction Brief up A short quiz Demo Lecture...

Activity Scheduled Action/ timing

Teaching Assistant Introduction Brief up A short quiz Demo Lecture by Prof Li. Chapter 3

2 minutes10 minutes Purifying, detecting, and Characterizing Proteins (p92)

Videos- SDS Gel filtration 3 minutes

Lecture Continued

Video– Gel, Affinity and ion exchange chromatography

8 minutes

Assignment- Review chapter/section covered in class,

Quiz in next class, 5-10 minutes

Class test Chapter 3 (Analyze the data ) Scheduled on 24 March, 2013

Teaching Plan- Chapter 5 March 19, 2015

Important Information

• Evaluation :

60 % Class Performance Attendance Class interaction Class tests Group Performance Presentation skills Quiz

40 % Final Exams

Tests are important so that we can grade you. Easy scoring Better understanding

Its very important to let us know your concerns or queries regarding the class and the subject Suggestions are most welcome and appreciated !!!

Lodish Website details

• http://bcs.whfreeman.com/lodish7e

http://bcs.whfreeman.com/lodish7e

3.6 Purifying, detecting, and Characterizing Proteins (p92)

Differential Centrifugation

EXPERIMENTAL FIGURE 3-34Centrifugation techniques separate paticles that differ in mass or density

Rate-Zonal Centrifugation

Electrophoresis

SDS-PAGE

EXPERIMENTAL FIGURE 3-35SDS-polyactrylamide gel electrophoresis(SDS-PAGE) separates proteins primarily on the basis of their masses

2-D Gel Electrophoresis

Page 95 EXPERIMENTAL FIGURE 3-36Two-dimensional gel electrophoresis separates proteins on the basis of charge and mass

EXPERIMENTAL FIGURE 3-36Two-dimensional gel electrophoresis separates proteins on the basis of charge and mass

Video

• SDS Gel Electrophoresis (Website ) • http://bcs.whfreeman.com/lodish7e/

#800911__811695__

Liquid Chromatography

EXPERIMENTAL FIGURE 3-37(a)Three commonly used liquid chromatographic techniques separate proteins on the basis of mass, charge, or affinity for a specific binding partner

EXPERIMENTAL FIGURE 3-37 (b)Three commonly used liquid chromatographic techniques separate proteins on the basis of mass, charge, or affinity for a specific binding partner

EXPERIMENTAL FIGURE 3-37(c)Three commonly used liquid chromatographic techniques separate proteins on the basis of mass, charge, or affinity for a specific binding partner

Video

• Gel Filtration Chromatography • Ion Exchange Chromatography• Affinity Chromatography

Western Blotting/Immunoblotting

EXPERIMENTAL FIGURE 3-38Western blotting (immunoblotting) combines several techniques to resolve and detect a specific protein

Video

• Western Blot http://bcs.whfreeman.com/lodish7e/#800911__811695__

http://bcs.whfreeman.com/lodish7e/

http://bcs.whfreeman.com/lodish7e/

Radio Labeling

Radioisotopes are indispensable tools for detecting biological molecules

Pulse-Chase labeling

EXPERIMENTAL FIGURE 3-39Pulse-chase experiments can track the pathway of protein modification or movement within cells

Labeling Experiments and Detection of Radiolabeled Molecules

Mass Spectrometry

EXPERIMENTAL FIGURE 3-40Molecular mass can be determined by matrix-assisted laser desorption/ionization time-of –flight (MALDI-TOF) mass spectrometry

Mass Spectrometry Can Determine the Mass and Sequence of Proteins

Electrospray Ionization –Ion Trap-Mass

EXPERIMENTAL FIGURE 3-41(a)Molecular mass of proteins and peptides can be determinded by electrospray ionization ion-trap mass specrometry

EXPERIMENTAL FIGURE 3-41(a)Molecular mass of proteins and peptides can be determinded by electrospray ionization ion-trap mass specrometry

LC-MS/MS can determine protein sequence

Determine 3-D structure by X-ray Crystallography

Analysis

EXPERIMENTAL FIGURE 3-42(a)X-ray crystallography provides diffraction data from which the three-dimensional structure of a protein can be determined

EXPERIMENTAL FIGURE 3-42(a)X-ray crystallography provides diffraction data from which the three-dimensional structure of a protein can be determined

3.7 Proteomics-Definition and Techniques

EXPERIMENTAL FIGURE 3-43LC-MS/MS is used to identify the proteins in a complex biological sample

Advanced Techniques in Mass Spectrometry Are Critical to Proteomic Analysis

Identify Proteins: Density Gradient Centrifugation and LC-MS/MS

EXPERIMENTAL FIGURE 3-44(a)Density-gradient centrifugation and LC-MS/MS can be used to identify many of the proteins in organilles

EXPERIMENTAL FIGURE 3-44(a)Density-gradient centrifugation and LC-MS/MS can be used to identify many of the proteins in organilles

KEY CONCEPTS OF SECTION 3.7Proteomics

Key words in chapter 3.6 and 3.7

Isoelectric point(pI) 等电点Liquid chromatography(LC) 液相色谱Western blottingRadioisotope 放射性同位素Pulse-chase 脉冲追踪Proteomics 蛋白质组学

1. Gel Electrophoresis

2. Microarray

3. Immunostaining

The principle and Application

Prepare Student PresentationsSelect 1 topic

KEY WORDS

Deoxyribonucleic acid(DNA)Ribonucleic acid(RNA)Messenger RNA(mRNA)Transfer RNA(tRNA)Ribosomal RNA(rRNA)TranscriptionGene expressionPurinePyrimidinePhosphodiester bondDouble helixBase paircomplementary

脱氧核糖核酸核糖核酸信使ＲＮＡ转运ＲＮＡ核糖体ＲＮＡ转录翻译嘌呤嘧啶磷酸二酯键双螺旋碱基互补

DenaturationNucleic acid hybridizaitonMicro RNAs(miRNAs)RNA polymeraseUpstreamPromoterTranscription factorPrimary transcriptGenomeOperonPrecursor mRNAs(pre-mRNAs)RNA splicingAlternative splicingIsoformAnticodonExtronIntron

变性核酸杂交小ＲＮＡＲＮＡ聚合酶上游启动子转录因子原始转录的 RNA基因组操纵子前体ｍＲＮＡ

ＲＮＡ剪接可变剪接异构体反义密码子外显子内含子

RibosomeGenetic codeReading frameAminoacyl-tRNAInitiation factors(IFs)Elongation factors(EFs)Release factors(RFs)GTPase superfamilyPolyribosomeHelicaseLeading strandLagging strandOkazaki fragmentCyclin-dependent kinaseRecombination

核糖体密码子阅读框氨酰ｔＲＮＡ起始因子延伸因子释放因子ＧＴＰ酶超家族多核糖体解旋酶前导链后随链冈崎片断ＣＤＫ重组

Point mutationMeiosisCrossing overVirionBacteriophageCapsidNucleocapsidEnvelopePlaque assayLysogenyRetrovirusReverse transcriptaseProvirusOnco-gene

点突变减数分裂交换病毒噬菌体衣壳核衣壳外壳噬菌斑试验溶原现象反转录病毒反转录酶前病毒致癌基因

Discussion: Answer Questions:

1. How to Preview and Review 2. How to Present

Homework:

1. Review Chapter 4 Concepts p161 (will be tested in Final)2. Analyzing the data p161-162 (These will be tested in Final)

3. Prepare quiz for Chapter 5

Next Tuesday – Class test Chapter 3 questions and (Analyze the data )Scheduled on 24 March, 2015

ActivityScheduled Action/ timing Teaching Assistant Introduction Brief up A short quiz Demo Lecture...

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