title: Pharmacological properties and pathophysiological significance of a Kunitz-type protease inhibitor (Rusvikunin-II) and its protein complex (Rusvikunin complex) purified from Daboia russelii russelii venom.
abstract: A 7.1 kDa basic peptide (Rusvikunin-II) was purified from a previously described protein complex (Rusvikunin complex, consists of Rusvikunin and Rusvikunin-II) of Daboia russelii russelii venom. The N-terminal sequence of Rusvikunin-II was found to be blocked, but peptide mass fingerprinting analysis indicated its identity as Kunitz-type basic protease inhibitor 2, previously reported from Russell's Viper venom. A tryptic peptide sequence of Rusvikunin-II containing the N-terminal sequence HDRPTFCNLFPESGR demonstrated significant sequence homology to venom basic protease inhibitors, Kunitz-type protease inhibitors and trypsin inhibitors. The secondary structure of Rusvikunin-II was dominated by β-sheets (60.4%), followed by random coil (38.2%), whereas α-helix (1.4%) contributes the least to its secondary structure. Both Rusvikunin-II and the Rusvikunin complex demonstrated dose-dependent anticoagulant activity; however, the anticoagulant potency of latter was found to be higher. Both inhibited the amidolytic activity of trypsin > plasmin >> FXa, fibrinogen clotting activity of thrombin, and, to a lesser extent, the prothrombin activation property of FXa; however, the inhibitory effect of the Rusvikunin complex was more pronounced. Neither Rusvikunin-II nor Rusvikunin complex inhibited the amidolytic activity of chymotrypsin and thrombin. Rusvikunin-II at 10 μg/ml was not cytotoxic to Colo-205, MCF-7 or 3T3 cancer cells; conversely, Rusvikunin complex showed ∼30% reduction of MCF-7 cells under identical experimental conditions. Rusvikunin-II (5.0 mg/kg body weight, i.p. injection) was not lethal to mice or House Geckos; nevertheless, it showed in vivo anticoagulant action in mice. However, the Rusvikunin complex (at 5.0 mg/kg) was toxic to NSA mice, but not to House Geckos, suggesting it has prey-specific toxicity. Rusvikunin complex-treated mice exhibited dyspnea and hind-limb paresis prior to death. The present study indicates that the Kunitz-type protein complex Rusvikunin from Russell's Viper venom significantly contributes to venom toxicity, and an important biological role in venoms appears to be facilitation of prey subjugation.
authors: Microbial Biotechnology and Protein Research Laboratory, Department of Molecular Biology and Biotechnology, Tezpur University, Tezpur 784 028, Assam, India; School of Biological Sciences, University of Northern Colorado, Greeley, CO 80639-0017, USA. Electronic address: akm@tezu.ernet.in.
authors: Ashis K
authors: AK
authors: Mukherjee
authors: School of Biological Sciences, University of Northern Colorado, Greeley, CO 80639-0017, USA.
authors: Stephen P
authors: SP
authors: Mackessy
mesh_terms: D000818:Animals; D000925:Anticoagulants / Q000737:chemistry* / Q000302:isolation & purification / Q000633:toxicity; D001412:Bacillus subtilis / Q000187:drug effects; D004926:Escherichia coli / Q000187:drug effects; D006041:Goats; D008116:Lizards; D051379:Mice; D008826:Microbial Sensitivity Tests; D017434:Protein Structure, Tertiary; D030162:Reptilian Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000633:toxicity; D015842:Serine Proteinase Inhibitors / Q000737:chemistry* / Q000302:isolation & purification / Q000633:toxicity; D014757:Viper Venoms / Q000737:chemistry*
chemical_list: D000925:Anticoagulants; D030162:Reptilian Proteins; D015842:Serine Proteinase Inhibitors; D014757:Viper Venoms
keywords: Biological role; Factor Xa inhibitor; Serine protease inhibitor; Thrombin inhibitor; Toxicity; Venom protein complex