A new method for C-terminal sequence analysis in the proteomic era.
The overall study of post-translational modifications (PTMs) of proteins is gaining strong interest. Beside phosphorylation and glycosylation, truncations of the nascent polypeptide chain at the amino or carboxy terminus are by far the most common types of PTMs in proteins. In contrast to the analysis of phosphorylation and glycosylation sites, relatively little attention has been paid to the development of approaches for the systematic analysis of proteolytic processing events. Here we present a new mass spectrometry (MS)-based strategy that allows the identification of the C-terminal sequence of proteins. The method can be directly applied to proteins cleaved with cyanogen bromide (CNBr) and purified either by SDS-PAGE, by two-dimensional (2D) PAGE or in solution, and it therefore eliminates the need for specific isolation of the C-terminal peptide. Using Shewanella oneidensis as a model system, we have demonstrated that this approach can be used for C-terminal sequence analysis at a proteomic scale. We also applied the method to study the C-terminal proteolytic processing of procardosin A.
Samyn
Bart
B
Department of Biochemistry, Physiology and Microbiology, Laboratory of Protein Biochemistry and Protein Engineering, Ghent University, K.L. Ledeganckstraat 35, B-9000 Ghent, Belgium. bart.samyn@ugent.be
Sergeant
Kjell
K
Castanheira
Pedro
P
Faro
Carlos
C
Van Beeumen
Jozef
J
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D020543:Proteome / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020539:Sequence Analysis, Protein / Q000379:methods*; D020592:Shewanella / Q000235:genetics / Q000378:metabolism*; D013728:Terminator Regions, Genetic / Q000235:genetics
D001426:Bacterial Proteins; D020543:Proteome