Glycosaminoglycans on fibroblasts accelerate thrombin inhibition by protease nexin-1.
Protease nexin-1 (PN-1) is a proteinase inhibitor that is secreted by human fibroblasts in culture. PN-1 inhibits certain regulatory serine proteinases by forming a covalent complex with the catalytic-site serine residue; the complex then binds to the cell surface and is internalized and degraded. The fibroblast surface was recently shown to accelerate the rate of complex-formation between PN-1 and thrombin. The present paper demonstrates that the accelerative activity is primarily due to cell-surface heparan sulphate, with a much smaller contribution from chondroitin sulphate. This conclusion is supported by the effects of purified glycosaminoglycans on the second-order rate constant for the inhibition of thrombin by PN-1. Also, treatment of 35SO4(2-)-labelled cells with heparitin sulphate lyase or chondroitin sulphate ABC lyase demonstrated two discrete pools of 35S-labelled glycosaminoglycans; subsequent treatment of plasma membranes with these glycosidases showed that heparitin sulphate lyase treatment abolished about 80% of the accelerative activity and chondroitin sulphate ABC lyase removed the remaining 20%. These results show that two components are responsible for the acceleration of PN-1-thrombin complex-formation by human fibroblasts. Although dermatan sulphate is also present on fibroblasts, it did not accelerate the inhibition of thrombin by PN-1.
D016564:Amyloid beta-Protein Precursor; D002352:Carrier Proteins / Q000494:pharmacology*; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D002810:Chondroitinases and Chondroitin Lyases / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D006025:Glycosaminoglycans / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D011133:Polysaccharide-Lyases / Q000378:metabolism; D058255:Protease Nexins; D011956:Receptors, Cell Surface; D058258:Serpin E2; D013917:Thrombin / Q000037:antagonists & inhibitors*
D016564:Amyloid beta-Protein Precursor; D002352:Carrier Proteins; D006025:Glycosaminoglycans; D058255:Protease Nexins; D011956:Receptors, Cell Surface; C545486:SERPINE2 protein, human; D058258:Serpin E2; D006026:Glycoside Hydrolases; D013917:Thrombin; D002810:Chondroitinases and Chondroitin Lyases; D011133:Polysaccharide-Lyases; C021349:heparitinsulfate lyase