Glyco-analytical multispecific proteolysis (Glyco-AMP): A simple method for detailed and quantitative glycoproteomic characterization

Serenus Hua, Chloe Y. Hu, Bum Jin Kim, Sarah M. Totten, Myung Jin Oh, Nayoung Yun, Charles C. Nwosu, Jong Shin Yoo, Carlito B Lebrilla, Hyun Joo An

Research output: Contribution to journalArticlepeer-review

37 Scopus citations


Despite recent advances, site-specific profiling of protein glycosylation remains a significant analytical challenge for conventional proteomic methodology. To alleviate the issue, we propose glyco-analytical multispecific proteolysis (Glyco-AMP) as a strategy for glycoproteomic characterization. Glyco-AMP consists of rapid, in-solution digestion of an analyte glycoprotein (or glycoprotein mixture) by a multispecific protease (or protease cocktail). Resulting glycopeptides are chromatographically separated by isomer-specific porous graphitized carbon nano-LC, quantified by high-resolution MS, and structurally elucidated by MS/MS. To demonstrate the consistency and customizability of Glyco-AMP methodology, the glyco-analytical performances of multispecific proteases subtilisin, pronase, and proteinase K were characterized in terms of quantitative accuracy, sensitivity, and digestion kinetics. Glyco-AMP was shown be effective on glycoprotein mixtures as well as glycoproteins with multiple glycosylation sites, providing detailed, quantitative, site- and structure-specific information about protein glycosylation.

Original languageEnglish (US)
Pages (from-to)4414-4423
Number of pages10
JournalJournal of Proteome Research
Issue number10
StatePublished - Oct 4 2013


  • biopharmaceutical glycoproteins
  • glycan isomers
  • glycoproteomics
  • multispecific proteases
  • nonspecific proteases
  • site-specific glycosylation

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)


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