Unique Degradation Of Fibrinogen By Western Diamondback Rattlesnake Venom Proteases

Abstract

The venom of western diamondback rattlesnake, Crotalus atrox, renders blood unclottable even after addition of thrombin. The phenomenon is due to a limited proteolytic degradation of fibrinogen. Purified fibrinogen (1 mg/ml) or human plasma incubated with crude venom (7.5 μg/ml) became unclottable. Fibrinogen in a purified system was affected at a faster rate than that in plasma suggesting that plasma protease inhibitors partially inhibited the proteolysis of fibrinogen. The degradation of plasma fibrinogen did not result from plasminogen activation since the venom did not activate purified human plasminogen as determined by an amidolytic assay and SDS polyacrylamide gel electrophoresis. The degradation of the fibrinogen molecule (Mr 340,000) proceeded steadily with a continuous cleavage of small peptides as demonstrated by SDS polyacrylamide gel electrophoresis. The bulk of the degraded fibrinogen was represented in gels as a single band with gradually increasing electrophoretic mobility as the incubation time progressed. The correlation of molecular weight and clottability demonstrated that the earliest unclottable derivative had Mr 285,000; its Aα and Bβ chains were degraded loosing peptides of Mr 20,000 and 6,500, respectively, but the γ chain appeared intact. The degradation pattern of fibrinogen in plasma was different than that of fibrinogen alone. An unclottable fibrinogen derivative isolated from plasma by precipitation with 2.2 M glycine had Mr 325,000; its Aα and γ chains appeared unaffected but the Bβ chain had lost a peptide of Mr 6,500. This product represents a novel unclottable derivative of fibrinogen with apparently intact Aα and γ chains, cleaved Bβ chain and high molecular weight. The cleavage of the Bβ chain would indicate that the intact Bβ chain may have an important role in the conversion of fibrinogen to fibrin clot.