LECTURE-01-PPT-v2 - NPTELnptel.ac.in/courses/102101007/downloads/PPT/LEC-01-PPT.pdf ·...

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Dr. Sanjeeva Srivastava IIT Bombay

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Genome

Transcriptome

Proteome

Splicing Post-translational modification (PTM)

IIT Bombay Proteomics Course NPTEL 2

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Protein Extraction

Protein Separation

Protein Identification

Protein Characterization


Proteomics

IIT Bombay Proteomics Course NPTEL 4 Curr Opin Chem Biol 2005, 9, 14-9

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Proteomics 2D Gel Electrophoresis

Mass spectrometry

Nucleotide sequencing ESTs/Genome scale

Algorithms Genetic approaches

Chip based approaches

1975 1996 1994 1990 2002

time

Complex mixture analysis (LC MS/MS)

IIT Bombay Proteomics Course NPTEL 5 Nature genetics 2003, 33, 311-323

IIT Bombay Proteomics Course NPTEL

•  Protein chemistry to proteomics

•  Diversification of proteomics technologies

•  Advent of novel technology platforms

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•  Gene expression regulated by post-transcriptional and post-translational modifications

•  Compared to genome of an organism proteome presents further complexity

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•  Orderly and unidirectional flow of information encoded in base sequences of cells, DNA to RNA and protein

•  Understanding biological system is a challenging task

•  Omics approaches are promising

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IPG strip

2-D ELECTROPHORESIS (2-DE)

SDS-PAGE

DIFFERENCE GEL ELECTROPHORESIS

(DIGE)

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Dass et al. DOI: 10.1002/9780470118498.ch3


Ioniza'on Mass Analyzer Detec'on

Detects ions

Forms ions

(charged molecules)

Sort Ions by Mass (m/z)

Data Processing

Rela=ve

Abu

ndance

m/z 1000 2000

Mass Spectrum

Data System

Ion Source

Ion Detector

Sample Inlet

Mass Analyzer

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Ramachandran et al. 2005. FEBS J; Chandra et al. 2010. Proteomics


Functionalized array surface

Heterologous host (e.g. E. coli)

Gene of interest

Gene insert

Protein expression

Protein purification Expression

vector

Chromatography

Protein purification SDS-PAGE

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Bound antibodies Flow cell system

Gold film

Change in angle of reflection

Free antigen

Glass slide

Antigen-Antibody complex

Prism

IIT Bombay Proteomics Course NPTEL 18 Ray et al. 2010. Proteomics

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•  Unraveling structural and functional details of proteins at the proteome level is a daunting task

•  Proteomics has quickly evolved to become an integral aspect of human biology and medicine

•  Proteomics has advanced rapidly; however, many experimental and computational challenges still exist

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•  Introduction to Proteomics: Tools for the New Biology, D.C. Liebler, Humana Press, 2002.

•  Principles of Proteomics, R.M.Twyman, BiosScientific Pub.,2004.

•  Proteomics for Biological Discovery, T.D.Veenstra, J.R.YatesIII, John- Wiley & Sons, Hoboken, New Jersey, USA; 2006.

•  Protein Biochemistry and Proteomics (The Experimenter Series), R. Hubert, Academic Press, 2006.

•  Proteomics in Practice: A Guide to Successful Experimental Design, R. Westermeier, T. Naven, H-R. Höpker, Wiley-VCH, 2008.

•  Curr Opin Chem Biol. 2005 Feb;9(1):14-9. Protein microarrays as tools for functional proteomics. LaBaer J, Ramachandran N.

•  Nature genetics 2003, 33, 311-323. Proteomics: the first decade and beyond - Nature Genetics. Scott D. Patterson & Ruedi H. Aebersold.

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•  Ray et al., 2010. Label-free detection techniques for protein microarrays: Prospects, merits and challenges, Proteomics. Volume 10, Issue 4, pages 731–748, No. 4.

•  Ray et al. 2012. Virtual Labs in proteomics: New E-learning tools. J of Proteomics. Volume 75, Issue 9, 17 May 2012, Pages 2515–2525

•  Chandra et al. 2010. Cell-free synthesis-based protein microarrays and their applications. Proteomics. Volume 10, Issue 4, pages 717–730, No. 4 February 2010.

•  Ramachandran et al. 2005. Emerging tools for real-time label-free detection of interactions on functional protein microarrays. FEBS J. Volume 272, Issue 21, pages 5412–5425, November 2005.

•  Qiang Tian1, W. Andy Tao. Chemical and Functional Genomic Approaches to Stem Cell Biology and Regenerative Medicine. Chapter 10. Proteomics in Stem Cells. ISBN: 9780470041468


•  Dass C. Chapter 3. Mass Analysis and Ion Detection. Fundamentals of Contemporary Mass Spectrometry. DOI: 10.1002/9780470118498.ch3.

•  Arthur H. J Clin Invest. Protein aggregation in disease: a role for folding intermediates forming specific multimeric interactions. 2002;110(9):1221–1232. doi:10.1172/JCI16781. Volume 110, Issue 9 (November 1, 2002).

•  Ray et al. 2010. Nanotechniques in proteomics: Current status, promises and challenges. J of Proteomics. Volume 25, Issue 11, 15 July 2010, Pages 2389–2401.

•  Roden LD. 1996. Global Analysis of a Macromolecular Interaction Measured on BIAcore. Biochemical and Biophysical Research Communications. Volume 225, Issue 3, 23 August 1996, Pages 1073–1077

•  Dhingra et al. 2005. New frontiers in proteomics research: A perspective. International Journal of Pharmaceutics. Volume 299, Issues 1–2, 11 August 2005, Pages 1–18.

•  Homola et al. Present and future of surface plasmon resonance biosensors. Analytical and Bioanalytical Chemistry. October 2003, Volume 377, Issue 3, pp 528-539

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•  My mentors Profs. Nat Kav and Joshua LaBaer

•  All the guest professors and application experts for stimulating discussion

•  All students and support staff (Ms. Harini S., Ms. Renisa D., Dr. Parvez S., Dr. Jaya V., Mr. Saurabh Y., Mr. Khaja S., Mr. Sandipan R., Mr. Jaipal PR, Mr. Karthik K., Ms. Rekha J.) for their help and contributions in NPTEL proteomics course

•  CDEEP team (Mr. Arun K., Mr. Tushar D., Ms. Sangeeta S.)

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