Immunological characterization of human vitronectin and its binding to glycosaminoglycans.
The cell-adhesive glycoprotein vitronectin in human plasma was characterized with a monospecific anti-vitronectin antibody. Vitronectin, a mixture of monomeric 75 and 65 kDa polypeptides, was found to have different ratios of amounts of 75 and 65 kDa polypeptides in immunoblots of sera from various healthy human donors. Two states of vitronectin were previously reported; the open state binds to heparin, but the cryptic state does not (Hayashi et al. (1985) J. Biochem. 98, 1135-1138). The anti-vitronectin antibody was suggested to react more strongly with the open state of vitronectin than with the cryptic state. To quantitate all vitronectin regardless of its state, an enzyme-linked immunosorbent assay of vitronectin was developed based on prior boiling of vitronectin-containing samples in 2% (w/v) sodium dodecyl sulfate and 40 mM dithiothreitol to destroy conformational differences. About 12-20% of the vitronectin molecules in plasma were found to bind to heparin-Sepharose under physiological conditions. Vitronectin in plasma bound 30-fold more efficiently to heparin immobilized by amino groups than by carboxyl groups. Its affinity for heparin was higher than for chondroitin sulfate A or C, or dermatan sulfate. Vitronectin was also found to contain covalently-linked small polypeptides of 15 and 13 kDa. These light chains seemed to be disulfide-bonded to the 65 kDa polypeptide, and might be endogenously derived from nicks in the carboxy-terminal portion of the 75 kDa polypeptide in plasma.
D002845:Chromatography; D004797:Enzyme-Linked Immunosorbent Assay; D006023:Glycoproteins / Q000097:blood* / Q000276:immunology; D006025:Glycosaminoglycans / Q000378:metabolism*; D006493:Heparin / Q000378:metabolism; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D007159:Immunologic Tests; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification; D019096:Vitronectin
D006023:Glycoproteins; D006025:Glycosaminoglycans; D007106:Immune Sera; D010446:Peptide Fragments; D019096:Vitronectin; D006493:Heparin