N-glycans of Human Protein C Inhibitor: Tissue-Specific Expression and Function

Abstract

Protein C inhibitor (PCI) is a serpin type of serine protease inhibitor that is found in many tissues and fluids in human, including blood plasma, seminal plasma and urine. This inhibitor displays an unusually broad protease specificity compared with other serpins. Previous studies have shown that the N-glycan(s) and the NH2-terminus affect some blood-related functions of PCI. In this study, we have for the first time determined the N-glycan profile of seminal plasma PCI, by mass spectrometry. The N-glycan structures differed markedly compared with those of both blood-derived and urinary PCI, providing evidence that the N-glycans of PCI are expressed in a tissue-specific manner. The most abundant structure (m/z 2592.9) had a composition of Fuc3Hex5HexNAc4, consistent with a core fucosylated bi-antennary glycan with terminal Lewisx. A major serine protease in semen, prostate specific antigen (PSA), was used to evaluate the effects of N-glycans and the NH2-terminus on a PCI function related to the reproductive tract. Second-order rate constants for PSA inhibition by PCI were 4.3±0.2 and 4.1±0.5 M−1s−1 for the natural full-length PCI and a form lacking six amino acids at the NH2-terminus, respectively, whereas these constants were 4.8±0.1 and 29±7 M−1s−1 for the corresponding PNGase F-treated forms. The 7–8-fold higher rate constants obtained when both the N-glycans and the NH2-terminus had been removed suggest that these structures jointly affect the rate of PSA inhibition, presumably by together hindering conformational changes of PCI required to bind to the catalytic pocket of PSA.

Tumor Biomarker Glycoproteins in the Seminal Plasma of Healthy Human Males Are Endogenous Ligands for DC-SIGN

“…The level of this glycoprotein in seminal plasma is about 160 Ϯ 20 g/ml, or about 30-to 40-fold higher than in human plasma (65). PCI rapidly inactivates PSA after ejaculation (66). Complexes of PCI and PSA are present in seminal plasma, consistent with this proposed physiological role (65).…”

“…However, such complexes were not detected in the fractions bound to Lotus lectin-agarose. Recently, PCI was immunopurified from seminal plasma and its oligosaccharides subjected to structural analysis (66). The seminal plasma glycoforms of this glycoprotein expressed: (1) very low levels of high mannosetype oligosaccharides; (2) virtually no sialylated antennae; (3) an extremely high density of terminal Lewis x and Lewis y sequences on its complex type N-glycans; and (4) no O-glycans.…”

Ovarian cyst fluid is a rich proteome resource for detection of new tumor biomarkers

“…In blood PCI is found in various forms including inactive, cleaved, and complex-bound. PCI is also modified through post-translational events and has been reported as microheterogeneous as a result of differences in N-glycan structures [35]. …”

Site-specific Glycoforms of Haptoglobin in Liver Cirrhosis and Hepatocellular Carcinoma

“…Multiply fucosylated N-glycans were detected in seminal plasma (44). One of the proteins extensively fucosylated in seminal plasma is protein-C inhibitor, but this protein in plasma carries Nglycans with at most two fucoses (45,46). Zona pellucida and sperm N-glycans with extensive fucosylation were described to modify protein interactions in the reproductive context, although a recent report described hyper-fucosylated lactosamines with up to six fucoses on N-linked glycans in pancreatic cyst fluids primarily associated with ␣-amylase and triacylglycerol lipase (47,48).…”