De novo peptide sequencing and quantitative profiling of complex protein mixtures using mass-coded abundance tagging.
Proteomic studies require efficient, robust, and practical methods of characterizing proteins present in biological samples. Here we describe an integrated strategy for systematic proteome analysis based on differential guanidination of C-terminal lysine residues on tryptic peptides followed by capillary liquid chromatography-electrospray tandem mass spectrometry. The approach, termed mass-coded abundance tagging (MCAT), facilitates the automated, large-scale, and comprehensive de novo determination of peptide sequence and relative quantitation of proteins in biological samples in a single analysis. MCAT offers marked advantages as compared with previously described methods and is simple, economic, and effective when applied to complex proteomic mixtures. MCAT is used to identify proteins, including polymorphic variants, from complex mixtures and measure variation in protein levels from diverse cell types.
Cagney
Gerard
G
Program in Proteomics and Bioinformatics, Banting and Best Department of Medical Research, University of Toronto, Toronto, ON, Canada.
Emili
Andrew
A
D000465:Algorithms; D000595:Amino Acid Sequence; D001331:Automation; D002853:Chromatography, Liquid / Q000379:methods; D005656:Fungal Proteins / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D020539:Sequence Analysis, Protein / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D013997:Time Factors; D014357:Trypsin / Q000737:chemistry
D005656:Fungal Proteins; D010455:Peptides; D011506:Proteins; D014357:Trypsin; D008239:Lysine