Activation of proPHBSP, the zymogen of a plasma hyaluronan binding serine protease, by an intermolecular autocatalytic mechanism

Citation
M. Etscheid et al., Activation of proPHBSP, the zymogen of a plasma hyaluronan binding serine protease, by an intermolecular autocatalytic mechanism, BIOL CHEM, 381(12), 2000, pp. 1223-1231
Citations number
29
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Biochemistry & Biophysics
Journal title
BIOLOGICAL CHEMISTRY
ISSN journal
1431-6730 → ACNP
Volume
381
Issue
12
Year of publication
2000
Pages
1223 - 1231
Database
ISI
SICI code
1431-6730(200012)381:12<1223:AOPTZO>2.0.ZU;2-R
Abstract
The hyaluronic acid binding serine protease (PHBSP), an enzyme with the abi lity to activate the coagulation factor FVII and the plasminogen activator precursors and to inactivate factor VIII and factor V, could be isolated fr om human plasma in the presence of 6 M urea as a single-chain zymogen, wher eas under native conditions only its activated two-chain form was obtained. The total yield of proenzyme (proPHBSP) was 5-6 mg/l, corresponding to a c oncentration of at least 80-100 nM in plasma. Upon removal of urea, even in the absence of charged surfaces a rapid development of amidolytic activity was observed that correlated with the appearance of the two-chain enzyme. The highest activation rate was observed at pH 6. ProPHBSP processing was c oncentration-dependent following a second order kinetic and was accelerated by catalytic amounts of active PHBSP, indicating an intermolecular autocat alytic activation. Charged macromolecules like poly-L-lysine, heparin, and dextran sulfate strongly accelerated the autoactivation, suggesting that in vivo proPHBSP activation might be a surface-bound process. The intrinsic a ctivity of the proenzyme was determined to be 0.25-0.3%, most likely due to traces of PHBSP. The presence of physiological concentrations of known pla sma inhibitors of PHBSP, like alpha (2) antiplasmin and C1 esterase inhibit or, but not antithrombin III/heparin, slowed down zymogen processing. Our i n vitro data suggest that the autoactivation of proPHBSP during plasma frac tionation is induced by the removal of inhibitors of PHBSP and is accelerat ed by charged surfaces of the chromatographic resins.