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Proteome Analysis

Proteome Analysis. July 3, 2007 Learning objectives. Understand the theoretical basis of 2D-SDS Gel Electrophoresis. Understand the latest technique for identifying proteins using mass spectrometry. Become familiar with the SWISS-2DPAGE Database

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Proteome Analysis

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  1. Proteome Analysis • July 3, 2007 • Learning objectives. Understand the theoretical basis of 2D-SDS Gel Electrophoresis. Understand the latest technique for identifying proteins using mass spectrometry. Become familiar with the SWISS-2DPAGE Database • Workshop on identifying proteins with the pI, MW and a few peptide masses.

  2. What is the proteome? metabolome proteome transcriptome genome

  3. 2D-SDS Gel ElectrophoresisWhat is it? • This is a method for the separation and identification of proteins in a sample by displacement in 2 dimensions oriented at right angles to one another. • Proteins separate over a large area, having a greater chance of resolution.

  4. First dimension • Separation by isoelectric point • Isoelectric point-pH at which the net charge on the molecule is zero

  5. Visuals http://www.aber.ac.uk/parasitology/Proteome /Tut_2D.html#Section%203

  6. Figure 4-44. Alberts et al. Molecular Biology of the Cell, 3rd edn.

  7. 2nd Dimension-to separate by size • Five steps: 1. Isolate proteins 2. Denature proteins 3. Load onto gel 4. Apply voltage gradient 5. Stain

  8. http://www.davidson.edu/academic/biology /courses/Molbio/SDSPAGE/SDSPAGE.html

  9. http://www.davidson.edu/academic/biology /courses/Molbio/SDSPAGE/SDSPAGE.html

  10. http://www.davidson.edu/academic/biology /courses/Molbio/SDSPAGE/SDSPAGE.html

  11. Figure 4-44. Alberts et al. Molecular Biology of the Cell, 3rd edn.

  12. Figure 3-42. Lodish et al. Molecular Cell Biology.

  13. Databases of 2D-Gels http://www-lmmb.ncifcrf.gov/EP/table2Ddatabases.html http://www.umh.ac.be/~biochim/balhf.htm

  14. http://www.weihenstephan.de/blm/deg/ manual/manualwork2html02testp9.htm

  15. Detector Reflection Mode Plate Laser Linear Mode 975.43 Detector Ab % Vacuum m/z

  16. http://www.matrixscience.com/cgi/search_form.pl?FORMVER=2&SEARCH=MIShttp://www.matrixscience.com/cgi/search_form.pl?FORMVER=2&SEARCH=MIS

  17. Protein Prospector • Another Web-based program is Protein Prospector (UCSF) • http://prospector.ucsf.edu/

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