Proteomics sequence tags

Powell, M. J., Sutton, J. N., Del Castillo, C. E., and Timperman, A. I., Marine proteomics Generation of sequence tags for dissolved proteins in seawater using tandem mass spectrometry. Marine Chemistry 95(3 ), 183-198, 2005. 

Fig. 17.9. Schematic diagram of MuDPIT proteomics (adapted from Yates, 1998). Protein mixtures of interest are cleaved via proteolytic activity and separated via a series of liquid chromatography separations. Identification of the proteins present relies on sequence tags produced from tandem mass spectrometry. Even low-abundance proteins should be represented at least on one occasion.

The study of the proteome of the recombinant adenovirus type 5 vectors demonstrated an important apphcation of separation techniques in combination with MS methods in the drug discovery process. With completely sequenced adenovirus genome available, this approach provides a chemically well-dehned method of characterization of structural proteins of recombinant adenoviral vectors. The information of protein MWs, tryptic peptide mass mapping, and sequence tags of tryptic peptides derived from HPLC/MS resulted in the identification of 17 adenoviral proteins/polypeptides in the purified virion. The rapid and accurate identification of viral proteins from recombinant adenoviruses in this study is significant since it provides direct evidence of the maturation stage of adenoviruses, which is closely related to viral infectivity and efficacy in gene therapy. 

A sequence tag from an unknown protein allows a number of further options for characterisation. Even a short stretch of amino acid sequence provides a powerful means of interrogating a protein database, and may provide a useful alternative to PMF in poor quality samples that may have peptides derived from more than one protein. More advanced database searching systems will find proteins with homologous sequences to that of the peptide tag. A good example of this type of database searching system is BLAST (Basic Local Alignment Sequence Tool), (Altschul et al., 1997). A modified form of this (MS BLAST) was used in conjunction with PMF to characterise the proteome of the yeast Pichia pastoris, whose genome has not been fully sequenced... 

Some bioinformatics software tools for proteomics combine data analysis, statistics and artificial intelligence methods to manage MS data, to identify proteins and to update databases. In this section, specific tools used to identify proteins are reviewed. They use lists of peptide mass values from MS or MS/MS as input, and they may also combine this information with amino acid sequence tag information or amino acid composition to enhance the identification of proteins. Figure 6 shows a simplified flow chart of sample preparation and MS data collection. It also shows the techniques and tools for protein identification described in this section. 

Kuhn, K. Prinz, T. Schafer, J. Baumann, C. Scharfke, M. Kienle, S. Schwarz, J. Steiner, S. Hamon, C. Protein sequence tags a novel solution for comparative proteomics. Proteomics 2005, 5, 2364—2368. 

Savitski, M.M. Nielsen, M.L. Zubarev, R.A. New database-independent, sequence tag-based scoring of peptide MS/MS data validates Mowse scores, recovers below threshold data, singles out modified peptides and assesses the quahty of MS/MS techniques. Mol. Cell. Proteomics 2005, 4, 1180-1188. 

Peptide sequence tags with multidimensional protein identification technology (MudPIT). MudPIT is a gel-free shotgun proteomic approach, which does... 

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