title: Purification and characterization of multisquamase, the prothrombin activator present in Echis multisquamatus venom.
abstract: The venom of Echis multisquamatus (Central Asian sand viper) contains a single prothrombin activator, designated multisquamase, which is structurally and functionally different from ecarin, the prothrombin activator from the venom of Echis carinatus (saw-scaled viper). Multisquamase is comprised of a 58000 Mr and a 23000 Mr subunit that consists of two disulfide-linked chains of 12000 Mr and 10000 Mr, respectively. In contrast to ecarin, which activates prothrombin and prethrombin 1 at comparable rates, and whose activity is hardly affected by Ca2+ or by changes in ionic strength, multisquamase hardly activates prethrombin 1; prothrombin activation requires Ca2+ and is strongly inhibited at high ionic strength. The most favourable kinetic parameters are observed at 1 mM Ca2+ and at low ionic strength (Km=0.085 microM and kcat=0.68 s(-1) at I approximately 0.04). An increase in ionic strength considerably reduces the rate of prothrombin activation, due to an increase of the Km (Km=0.8 microM and kcat=1.03 s(-1) at I approximately 0.2). Studies in plasmas from patients on oral anticoagulant therapy show that E. Multisquamatus venom only activates carboxylated prothrombin, whereas E. carinatus activates both prothrombin and descarboxyprothrombin. Thus, multisquamase-dependent prothrombin activation appears to require post-translational modification of the gla-domain. This venom prothrombin activator may, therefore, become a useful tool to quantitate prothrombin and descarboxyprothrombin in cases where vitamin K-dependent carboxylation of prothrombin is impaired.
authors.affiliation: Cardiovascular Research Institute Maastricht, Department of Biochemistry, Maastricht University, The Netherlands.
authors.forename: R J
authors.initials: RJ
authors.lastname: Petrovan
authors.forename: J W
authors.initials: JW
authors.lastname: Govers-Riemslag
authors.forename: G
authors.initials: G
authors.lastname: Nowak
authors.forename: H C
authors.initials: HC
authors.lastname: Hemker
authors.forename: J
authors.initials: J
authors.lastname: Rosing
authors.forename: G
authors.initials: G
authors.lastname: Tans
mesh_terms: D004798:Enzymes / Q000302:isolation & purification / Q000378:metabolism; D008666:Metalloendopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D011516:Prothrombin / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism*; D014757:Viper Venoms / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D004798:Enzymes; D014757:Viper Venoms; D011516:Prothrombin; D013917:Thrombin; D008666:Metalloendopeptidases; C103417:ecamulin