N-glycosylation microheterogeneity and site occupancy of an Asn-X-Cys sequon in plasma-derived and recombinant protein C

Research output: Contribution to journalArticle

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Abstract

Human protein C (hPC) is glycosylated at three Asn-X-Ser/Thr and one atypical Asn-X-Cys sequons. We have characterized the micro- and macro-heterogeneity of plasma-derived hPC and compared the glycosylation features with recombinant protein C (tg-PC) produced in a trans- genic pig bioreactor from two animals having approximately tenfold different expression levels. The N-glycans of hPC are complex di- and tri-sialylated structures, and we measured 78% site occupancy at Asn-329 (the Asn-X-Cys sequon). The N-glycans of tg-PC are complex sialylated structures, but less branched and partially sialylated. The porcine mammary epithelial cells gly- cosylate the Asn-X-Cys sequon with a similar efficiency as human hepatocytes even at these high expression levels, and site occupancy at this sequon was not affected by expression level. A distinct bias for particular structures was present at each ofthe four glycosylation sites for both hPC and tg-PC. Interestingly, glycans with GalNAc in the antennae were predominant at the Asn-329 site. The N-glycan structures found for tg-PC are very similar to those reported for a recombinant Factor IX produced in transgenic pig milk, and similar to the endogenous milk protein lacto- ferrin, which may indicate that N-glycan processing in the porcine mammary epithelial cells is more uniform than in other tissues.

Original languageEnglish (US)
Pages (from-to)2555-2567
Number of pages13
JournalProteomics
Volume9
Issue number9
DOIs
StatePublished - May 1 2009

Fingerprint

Glycosylation
Protein C
Recombinant Proteins
Polysaccharides
Blood Proteins
Plasmas
Swine
Breast
Epithelial Cells
Factor IX
Lactoferrin
Milk Proteins
Bioreactors
Macros
Hepatocytes
Milk
Animals
Tissue
Antennas
Processing

Keywords

  • Asn-X-Cys
  • Glycosylation
  • Mass spectrometry
  • N-glycan
  • Protein C

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

Cite this

N-glycosylation microheterogeneity and site occupancy of an Asn-X-Cys sequon in plasma-derived and recombinant protein C. / Gil, Geun Cheol; Velander, William H.; Van Cott, Kevin E.

In: Proteomics, Vol. 9, No. 9, 01.05.2009, p. 2555-2567.

Research output: Contribution to journalArticle

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