Peptide cleavage sites identification

Technologies that offer the opportunity to use MS for intact protein mass determination allow rapid assessment of signal peptide cleavage sites and post-translational heterogeneity. A well-resolved mass spectrum of an intact protein defines its native covalent profile as well as its associated heterogeneity. A comprehensive mass spectrometric approach that integrates both intact protein molecular mass measurement (top-down) and proteolytic fragment identification (bottom-up) would represent a powerful approach for proteome/ protein characterization. 

Nielsen, H., Engelbrecht, J., Brunak, S., and von Heijne, G. (1997). Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Protein Eng. 10, 1-6. 

Signal peptide identification, like DNA intron/exon sequence discrimination, involves the two related problems of signal peptide discrimination (search for content) and cleavage site recognition (search for signal). It is well suited to neural network methods for several reasons. The functional units are encoded by local, linear sequences of amino acids rather than global 3-dimensional structures (Claros et al., 1997). The ambiguity of... 

PS—alkyne probe, followed by a CC reaction to introduce a biotin tag with a tobacco etch virus (TEV) protease cleavage site. Tagged proteins were then enriched with streptavidin beads and subsequendy digested by trypsin. The supernatant contained peptide fragments of the enriched proteins and was, in contrast to the previous procedure, saved for subsequent analysis by multidimensional protein identification technology (MudPIT), a novel two-dimensional LC—MS/MS analysis method that will be discussed in more detail... 

G. (1997) Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites, Protein Engineering 10 1-6. 

Nielsen, H., J. Engelbrecht, S. Brunak, and G. von Heijne. 1997. A neural network method for identification of prokaryotic and eukaroytoic signal peptides and prediction of their cleavage sites. Internationl Journal of Neural Systems 8 581-99. 

Identification of different cleavage sites within the peptides The determination of the presence and relative location of a specific residue in a peptide, either secondary digestion of the primary digest with an enzyme of differing specificity or parallel digestions of aliquots of the same sample with enzymes of different specificity can be employed. 

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