Purification and characterization of an antithrombin III inactivating enzyme from the venom of the African night adder (Causus rhombeatus).
A serine proteinase was isolated from the venom of the night adder (Causus rhombeatus) by fast protein liquid chromatography (anion-exchange, gel filtration and hydrophobic interaction). The protein (termed CR-serpinase) had an estimated mol. wt of 45,500 as determined by SDS-PAGE, pI of 4.7 and a carbohydrate content of 18.9%. Incubation of CR-serpinase with purified human antithrombin III at a molar ratio of 1:66 resulted in a loss of more than 90% of the initial AT III activity within 10 min. The reaction was dependent on heparin. In SDS-PAGE inactivation of human antithrombin III was correlated with the occurrence of two cleavage products. The cleavage site in the antithrombin III molecule was determined to be Arg 393-Ser 394 by amino-terminal sequencing. CR-Serpinase had no thrombin-like activity since no fibrinogen conversion was induced and had no procoagulant activity. CR-Serpinase activity was not inhibited by antithrombin III-heparin and was not decreased by a 10-min preincubation in normal human plasma. Inactivation of antithrombin III by CR-serpinase appeared to be very specific.
Janssen
M
M
Pentapharm Ltd, CH-4002 Basle, Switzerland.
Meier
J
J
Freyvogel
T A
TA
D000595:Amino Acid Sequence; D000818:Animals; D000990:Antithrombin III / Q000037:antagonists & inhibitors*; D002241:Carbohydrates / Q000032:analysis; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D005342:Fibrinolysis / Q000187:drug effects; D006168:Guinea Pigs; D066298:In Vitro Techniques; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D010974:Platelet Aggregation / Q000187:drug effects; D012697:Serine Endopeptidases / Q000302:isolation & purification*; D014757:Viper Venoms / Q000201:enzymology* / Q000633:toxicity
D002241:Carbohydrates; D014757:Viper Venoms; D000990:Antithrombin III; C078561:CR-serpinase; D012697:Serine Endopeptidases