EFFECT OF CARBOHYDRATE SIDE-CHAIN OF TISSUE-TYPE PLASMINOGEN-ACTIVATOR ON ITS INTERACTION WITH PLASMINOGEN-ACTIVATOR INHIBITOR-1

Tissue-type plasminogen activator (t-PA) has a variable N-glycosylatio n processing in different cell lines. We studied an effect of the carb ohydrate side chain of t-PA molecule on the interaction of t-PA with p lasminogen activator inhibitor-1 (PAI-1), First, we analysed the carbo hydrate composition of t-PAs derived from human diploid fibroblasts (f ibroblast t-PA) and Bowes melanoma cells (melanoma t-PA). Per 1 mol of protein fibroblast t-PA possesses 2.2 mol of sialic acid and 7.7 mol of mannose, whereas melanoma t-PA had 0.9 mol of sialic acid and 9.7 m ot of mannose. The second-order rate constants (pH 7.4, 37 degrees C) for the interactions between type I or type II single-chain t-PA and P AI-1 were determined after fractionation of t-PA by lysine-Sepharose c hromatography. The rate constants of type I and type II single-chain f ibroblast t-PA were significantly different (P< 0.03). type 1-1.43 +/- 0.13 x 10(6) and type II-1.70 +/- 0.05 x 10(6) M-1S-1 respectively, w hereas those of melanoma t-PA were almost similar. Sialidase treatment of type I single-chain fibroblast t-PA apparently increased the rate constant, 1.68 +/- 0.10 x 10(6) M-1S-1, in almost an equal way to that of type Il t-PA. On the other hand, modification of high mannose type oligosaccharide with endoglycosidase H treatment did not alter the ra te constants of all types of t-PAs. These results suggest that the sia lic acid of N-glycosylation at the site 184 on single-chain t-PA modul ates the interaction with PAI-1.