Robust analysis of the yeast proteome under 50 kDa by molecular-mass-based fractionation and top-down mass spectrometry.

Kellie JF, Catherman AD, Durbin KR, Tran JC, Tipton JD, Norris JL, Witkowski CE, Thomas PM, Kelleher NL
Anal Chem. 2012 84 (1): 209-15

PMID: 22103811 · PMCID: PMC3262231 · DOI:10.1021/ac202384v

As the process of top-down mass spectrometry continues to mature, we benchmark the next installment of an improving methodology that incorporates a tube-gel electrophoresis (TGE) device to separate intact proteins by molecular mass. Top-down proteomics is accomplished in a robust fashion to yield the identification of hundreds of unique proteins, many of which correspond to multiple protein forms. The TGE platform separates 0-50 kDa proteins extracted from the yeast proteome into 12 fractions prior to automated nanocapillary LC-MS/MS in technical triplicate. The process may be completed in less than 72 h. From this study, 530 unique proteins and 1103 distinct protein species were identified and characterized, thus representing the highest coverage to date of the Saccharomyces cerevisiae proteome using top-down proteomics. The work signifies a significant step in the maturation of proteomics based on direct measurement and fragmentation of intact proteins.

© 2011 American Chemical Society

MeSH Terms (7)

Chromatography, Liquid Electrophoresis, Polyacrylamide Gel Mass Spectrometry Molecular Weight Proteome Saccharomyces cerevisiae Proteins Tandem Mass Spectrometry

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