Proteomics Elements

Objectives

It is intended that students know the main techniques used in separation, analysis and identification of proteins obtained from complex samples and in a health context. It is expected that they acquire the knowledge on the main challenges facing proteomics in health and to recognize the importance of this approach.

Program

1. Proteomics importance on a health context

Functional proteomics approach; Identify disease markers; Identifying pathologies through quantitative proteomic analysis;

2. Challenges to proteomics on a health context

Identification of proteins in a biological complex; Difficulties in the detection of minority proteins; Conjugation of techniques for separation and purification;

3. Separation and detection techniques

Sample preparation and handling; Chromatographic separation (HPLC, FPLC); Separation by electrophoresis (SDS-PAGE, native-dimensional, SDS-PAGE 2DE); Staining (Coomassie, colloidal Coomassie, silver, fluorescence, immunoassay);

4. Protein analysis techniques

Image-analysis (2D ImageMaster, Quantity one)

5. Protein identification techniques

Edman sequencing of the N-terminus; Mass spectrometry (MALDI-TOF, MALDI-TOF-TOF, ESI-MS-MS)

6. Analysis in Sillico

Databases (NCBI, Swiss Prot, MSDB, APIASTER); molecular analysis tools.

Teaching Methodologies

It is intended that the classes are composed of a presentation of some fundamental theoretical aspects, allowing to launch questions, encouraging students to participate actively in order to correlate the topics covered. In the second part, practical problems are presented to students to be solved. Bibliography will be indicated where students can consolidate and expand aspects related to the themes. WEB open-source tools will be provided for students to solve problems and to achieve autonomous and rational information.

Bibliography

McDonald WH, Yates JR 3rd. Curr Opin Mol Ther. 2003 Jun;5(3):302-9.

Colinge J, Magnin J, Dessingy T, Giron M, Masselot A. Proteomics. 2003 Aug;3(8):1434-40.

Lin D, Tabb DL, Yates JR 3rd. Biochim Biophys Acta. 2003 Mar 21;1646(1-2):1-10.

Hunter TC, Andon NL, Koller A, Yates JR, Haynes PA. J Chromatogr B Analyt Technol Biomed Life Sci. 2002 Dec 25;782(1-2):165-81.

McDonald WH, Yates JR 3rd. Dis Markers. 2002;18(2):99-105. Review.

Henzel WJ, Watanabe C, Stults JT. Protein identification: the origins of peptide mass fingerprinting. J Am Soc Mass Spectrom. 2003 Sep;14(9):931-42.

Yates JR 3rd, Eng JK, McCormack AL, Schieltz D., Method to correlate tandem mass spectra of modified peptides to amino acid sequences in the protein database. Anal Chem. 1995 Apr 15;67(8):1426-36.

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