Talk:Bottom-up proteomics

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MagnusPalmblad (talk) 11:33, 24 April 2008 (UTC) Is "bottom-up proteomics" not used in a much wider sense than for the identification of spots from a 2D gel as presently stated in the article? Almost any method which measures peptides or their sequences (by MS/MS) to infer identity or quantity of proteins is described as "bottom-up" in the literature. This includes MudPIT and similar approaches. What is the difference between "bottom-up" and "shotgun" proteomics? If there isn't any, perhaps these pages should be merged? --MagnusPalmblad

I have seen shotgun presented as both a subset and separate from bottom-up; I'm not sure if there is a broad consensus. My suggestion would be to bring both this article and shotgun proteomics up to B class and see where we are. If you have some good references, you can put them in using the Pubmed ID and template filler.[1] Diagram(s) would be good, too. An overview should go in protein mass spectrometry and proteomics itself has been needing some work for a while. Much to do here. --Kkmurray (talk) 14:57, 24 April 2008 (UTC)

How about something like this: "Bottom-up proteomics is a common method to identify proteins and characterize their amino acid sequences and post-translational modifications by proteolytic digestion of proteins prior to analysis by mass spectrometry. The proteins may first be purified by a method such as gel electrophoresis resulting in one or a few proteins in each proteolytic digest. Alternatively, the crude protein extract is digested directly, followed by one or more dimensions of separation of the peptides by liquid chromatography coupled to mass spectrometry, a technique known as shotgun proteomics. By comparing the masses of the proteolytic peptides or their tandem mass spectra with those predicted from a sequence database, peptides can be identified and multiple peptide identifications assembled into a protein identification." For the bibliography: Chait BT (October 2006). "Chemistry. Mass spectrometry: bottom-up or top-down?". Science (journal) 314 (5796): 65–6. doi:10.1126/science.1133987. PMID 17023639.  and Aebersold R, Mann M (March 2003). "Mass spectrometry-based proteomics". Nature 422 (6928): 198–207. doi:10.1038/nature01511. PMID 12634793.  From this it should be clear that shotgun proteomics is a subset of bottom-up proteomics. I'll see if I can make a diagram comparing bottom-up and top-down. --MagnusPalmblad (talk) 12:08, 25 April 2008 (UTC)

The analogy with DNA sequencing may be more confusing than enlightening. In de novo shotgun (or bottom-up) DNA sequencing, there is no template like a sequence database (the product of the sequencing). Instead short but overlapping sequences are assembled to longer "contigs", assembled into genes and chromosomes. Use of overlapping sequences from proteolytic peptides is hardly ever used in proteomics, as it is very difficult to get sufficient sequence coverage to assemble the full protein sequence. But as it is at least a theoretical possibility, using DNA sequencing as an analogy to bottom-up proteomics could be confusing. --MagnusPalmblad (talk) 12:29, 25 April 2008 (UTC)

Here's another one that makes your point (if implicitly):VerBerkmoes NC, Bundy JL, Hauser L, et al (2002). "Integrating 'top-down" and "bottom-up" mass spectrometric approaches for proteomic analysis of Shewanella oneidensis". J. Proteome Res. 1 (3): 239–52. PMID 12645901. . This is the earliest mention of bottom-up I could find: Kelleher, Neil L.; Hong Y. Lin, Gary A. Valaskovic, David J. Aaserud, Einar K. Fridriksson, and Fred W. McLafferty (1999). ""Top down versus bottom up protein characterization by tandem high-resolution mass spectrometry"". J. Am. Chem. Soc 121 (4): 806 -812. doi:10.1021/ja973655h. . I'll try to track down a counter-example, but I think that you may be right that the general definition puts proteolysis approaches into the bottom-up category. --Kkmurray (talk) 14:52, 25 April 2008 (UTC)

I changed the entry according to the discussion above and added a figure. Please modify it if necessary (I am not a native speaker).--MagnusPalmblad (talk) 15:32, 28 April 2008 (UTC)