Purification and characterization of calobin II, a second type of thrombin-like enzyme from Agkistrodon caliginosus (Korean viper).
In our previous report, we purified and cloned the gene of a thrombin-like enzyme, calobin, from the venom of Agkistrodon caliginosus (Hahn, B.S., Yang, K.Y., Park, E.M., Chang, I.M., Kim, Y. S., 1996. Purification and molecular cloning of calobin, a thrombin-like enzyme from Agkistrodon caliginosus (Korean viper). J. Biochem. 119, 835-843.). During the purification of calobin, a second type of thrombin-like protease was found and it was purified using Affi-Gel Blue and Mono-S cation-exchange chromatography. It was identified as a serine protease with a molecular weight of 41, 000 on SDS-PAGE and its isoelectric point was determined to be 7.4 by isoelectric focusing. It showed little azocaseinolytic and fibrinolytic activity. However, this enzyme acted on fibrinogen to form fibrin with a specific activity of 7,587 NIH equivalent units and also exhibited arginine esterase activity. Amino acid sequencing of the N-terminal region established a primary structure composed of Val-Ile-Gly-Gly-Asp-Glu-Cys-Asn-Ile-Asn-Glu-His-Arg-Phe-Leu-Val-Ala-X -Tyr. This sequence was entirely consistent with that of calobin and showed a high homology with other thrombin-like enzymes, such as ancrod, batroxobin and gyroxin. Based on the biochemical properties such as molecular weight and isoelectric point, we can demonstrate a second thrombin-like protein showing a high potent clotting activity from the venom of Korean viper.
Cho
S Y
SY
Natrual Products Research Institute, Seoul National University, South Korea.
Hahn
B S
BS
Yang
K Y
KY
Kim
Y S
YS
D017836:Agkistrodon / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D005343:Fibrinolytic Agents / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D012697:Serine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D013917:Thrombin / Q000302:isolation & purification*; D014757:Viper Venoms / Q000302:isolation & purification*
D005343:Fibrinolytic Agents; D014757:Viper Venoms; D012697:Serine Endopeptidases; C104841:calobin; D013917:Thrombin