Enhanced mass spectrometric mapping of the human GalNAc-type O-glycoproteome with SimpleCells.

ABSTRACT: Characterizing protein GalNAc-type O-glycosylation has long been a major challenge, and as a result, our understanding of this glycoproteome is particularly poor. Recently, we presented a novel strategy for high throughput identification of O-GalNAc glycosites using zinc finger nuclease gene-engineered "SimpleCell" lines producing homogeneous truncated O-glycosylation. Total lysates of cells were trypsinized and subjected to lectin affinity chromatography enrichment, followed by identification of GalNAc O-glycopeptides by nLC-MS/MS, with electron transfer dissociation employed to specify sites of O-glycosylation. Here, we demonstrate a substantial improvement in the SimpleCell strategy by including an additional stage of lectin affinity chromatography on secreted glycoproteins from culture

SUBMITTER: Vakhrushev SY 

PROVIDER: S-EPMC3617340 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature