title: An N-glycosylation Analysis of Human Alpha-2-Macroglobulin Using an Integrated Approach.
abstract: Assignment of glycosylation sites and site microheterogeneity is of both biological and clinical significance. Herein, the detailed N-glycosylation pattern of human serum alpha-2-macroglobulin was studied using an integrative approach, including permethylation of N-glycans, collision induced dissociation (CID) and electron transfer dissociation (ETD) of chymotryptic N-glycopeptides, and partial deglycosylation of chymotryptic N-glycopeptides with endo-β-N-acetylglucosaminidase F3 (Endo F3). Three N-glycosylation sites were found to be occupied by four biantennary complex type N-glycans using N-glycan analysis and the ETD/CID method. Endo F3 assisted mass spectrometric analysis yielded five N-glycosylation sites with and without core fucosylation. In total, six out of eight potential N-glycosylation sites were identified using this approach. This integrative approach was performed using only 10 μL of human serum for both N-glycosylation site assignment and site microheterogeneity determination.
authors.affiliation: Department of Chemistry, The University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Zhenxin
authors.initials: Z
authors.lastname: Lin
authors.forename: Andy
authors.initials: A
authors.lastname: Lo
authors.forename: Diane M
authors.initials: DM
authors.lastname: Simeone
authors.forename: Mack T
authors.initials: MT
authors.lastname: Ruffin
authors.forename: David M
authors.initials: DM
authors.lastname: Lubman