U.S. Department of Energy

Pacific Northwest National Laboratory

Enrichment and analysis of nonenzymatically glycated peptides: boronate affinity chromatography coupled with electron-transfer dissociation mass spectrometry.

TitleEnrichment and analysis of nonenzymatically glycated peptides: boronate affinity chromatography coupled with electron-transfer dissociation mass spectrometry.
Publication TypeJournal Article
Year of Publication2007
AuthorsZhang Q, Tang N, Brock JWC, Mottaz HM, Ames JM, Baynes JW, Smith RD, Metz TO
JournalJ Proteome Res
KeywordsAmino Acid Sequence, Boronic Acids, Chromatography, Affinity, Electron Transport, Glucose, Glycopeptides, Glycosylation, Mass Spectrometry, Molecular Sequence Data, Proteomics
Abstract

Nonenzymatic glycation of peptides and proteins by d-glucose has important implications in the pathogenesis of diabetes mellitus, particularly in the development of diabetic complications. However, no effective high-throughput methods exist for identifying proteins containing this low-abundance post-translational modification in bottom-up proteomic studies. In this report, phenylboronate affinity chromatography was used in a two-step enrichment scheme to selectively isolate first glycated proteins and then glycated, tryptic peptides from human serum glycated in vitro. Enriched peptides were subsequently analyzed by alternating electron-transfer dissociation (ETD) and collision induced dissociation (CID) tandem mass spectrometry. ETD fragmentation mode permitted identification of a significantly higher number of glycated peptides (87.6% of all identified peptides) versus CID mode (17.0% of all identified peptides), when utilizing enrichment on first the protein and then the peptide level. This study illustrates that phenylboronate affinity chromatography coupled with LC-MS/MS and using ETD as the fragmentation mode is an efficient approach for analysis of glycated proteins and may have broad application in studies of diabetes mellitus.

DOI10.1021/pr070112q
Alternate JournalJ. Proteome Res.
PubMed ID17488106
PubMed Central IDPMC2587408
Grant List06564 / / Wellcome Trust / United Kingdom
5R21DK071283 / DK / NIDDK NIH HHS / United States
DK-19971 / DK / NIDDK NIH HHS / United States
P41 RR018522-05 / RR / NCRR NIH HHS / United States
R33 DK071283-03 / DK / NIDDK NIH HHS / United States
RR018522 / RR / NCRR NIH HHS / United States
| Pacific Northwest National Laboratory