Thrombin-like enzyme from the venom of Bitis gabonica. Purification, properties, and coagulant actions.
Gabonase, an enzyme which acts on fibrinogen and factor XIII in uniquely thrombin-like ways, was purified to electrophoretic homogeneity from the venom of Bitis gabonica. On sodium dodecyl sulfate-polyacrylamide electrophoresis, the reduced protein behaved as a single chain with Mr = 30,600. The enzyme contains 20.6% carbohydrate, no free sulfhydryl groups and hence, from amino acid analysis, five disulfide bonds. Its extinction coefficient (E1%1cm) at 280 nm is 9.6. Its pI is 5.3. Gabonase has an active serine residue, is inactivated by phenylmethanesulfonyl fluoride, and has an active histidine which reacts with the chloromethyl ketone of tosyl-L-lysine. Its NH2-terminal amino acid sequence (Val-Val-Gly-Gly-Ala-Glu-Cys-Lys-Ile-Asp-Gly-His-Arg-Cys-Leu-Ala-Leu-Leu -Tyr-) is homologous to the B chain of thrombin. The activity of the enzyme is stabilized by calcium ion. It exhibits strong N alpha-p-tosyl-L-arginine methyl esterase activity, hydrolyzes tripeptide nitroanilide derivatives weakly or not at all, and cleaves no peptide bonds in insulin, glucagon, or the S peptide of ribonuclease. Gabonase clots fibrinogen with a specific activity of 45 NIH thrombin-equivalent units/mg, releasing both fibrinopeptides A and B and showing substrate inhibition at fibrinogen concentrations of 3 mg/ml or greater. The enzyme also activates factor XIII. It is not inactivated by either heparin or hirudin.
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D010450:Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D005337:Fibrin / Q000378:metabolism; D005340:Fibrinogen / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis; D012697:Serine Endopeptidases; D013045:Species Specificity; D013379:Substrate Specificity; D013917:Thrombin; D014757:Viper Venoms / Q000302:isolation & purification*
D000596:Amino Acids; D010446:Peptide Fragments; D014757:Viper Venoms; D005337:Fibrin; D005340:Fibrinogen; D010450:Endopeptidases; D012697:Serine Endopeptidases; C049532:gabonase; D013917:Thrombin