L PresentationPROTEOMICS培训讲义.pptVIP

L529 - Presentation PROTEOMICS- Yogita Mantri -Arvind Gopu 11/10/2003 Introduction – What is Proteomics? “The identification, characterization and quantification of all proteins involved in a particular pathway, organelle, cell, tissue, organ or organism that can be studied in concert to provide accurate and comprehensive data about that system.” /prodef.html Central lesson from eukaryotic genome projects Evolutionary complexity is not primarily determined by increasing the number of genes, but by increasing variation on the level of the synthesized proteins. This is achieved by generating MULTIPLE proteins from only ONE gene e.g. by different combinations of exons by alternative splicing post-translational protein processing (e.g. cleavage of pro-peptides) post-translational protein modifications (e.g. acetylation, glycosylation) modified central dogma: DNA -- RNA -- protein(s) it is important to perform analyses on the level of gene PRODUCTS Key Key advantage of proteomics Researchers work on the level of gene products and deal with genes that are really expressed to give a detectable PRODUCT and are not just expressed“ which only says they produce a detectable mRNA but it is not clear whether there is a gene product or not. Key limitation of proteomics Usually, only a fraction of the proteins synthesized can be detected in a proteomics experiment, whereas the expression of ALL genes can be monitored in a whole-genome array experiment. Key prerequisite of proteomics A genome sequence for the investigated organism or at least a collection of many cDNA sequences is required. Experimental BackgroundMass Spectrometry What is Mass Spec? Analytical tool measuring molecular weight (MW) of sample Only picomolar concentrations required Within an accuracy of 0.01% of total weight of sample and within 5 ppm for small organic molecules For a Mr of 40 kDa, there is a 4 Da error This means it can detect amino acid substitutions / post-translational modificati