title: Ringhalexin from Hemachatus haemachatus: A novel inhibitor of extrinsic tenase complex.
abstract: Anticoagulant therapy is used for the prevention and treatment of thromboembolic disorders. Blood coagulation is initiated by the interaction of factor VIIa (FVIIa) with membrane-bound tissue factor (TF) to form the extrinsic tenase complex which activates FX to FXa. Thus, it is an important target for the development of novel anticoagulants. Here, we report the isolation and characterization of a novel anticoagulant ringhalexin from the venom of Hemachatus haemachatus (African Ringhals Cobra). Amino acid sequence of the protein indicates that it belongs to the three-finger toxin family and exhibits 94% identity to an uncharacterized Neurotoxin-like protein NTL2 from Naja atra. Ringhalexin inhibited FX activation by extrinsic tenase complex with an IC50 of 123.8 ± 9.54 nM. It is a mixed-type inhibitor with the kinetic constants, Ki and Ki' of 84.25 ± 3.53 nM and 152.5 ± 11.32 nM, respectively. Ringhalexin also exhibits a weak, irreversible neurotoxicity on chick biventer cervicis muscle preparations. Subsequently, the three-dimensional structure of ringhalexin was determined at 2.95 Å resolution. This study for the first time reports the structure of an anticoagulant three-finger toxin. Thus, ringhalexin is a potent inhibitor of the FX activation by extrinsic tenase complex and a weak, irreversible neurotoxin.
authors.affiliation: Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore 119260, Singapore.
authors.forename: Bhaskar
authors.initials: B
authors.lastname: Barnwal
authors.affiliation: Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore 119260, Singapore.
authors.forename: Chacko
authors.initials: C
authors.lastname: Jobichen
authors.affiliation: Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore 119260, Singapore.
authors.forename: Vallerinteavide Mavelli
authors.initials: VM
authors.lastname: Girish
authors.affiliation: Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore 119260, Singapore.
authors.forename: Chun Shin
authors.initials: CS
authors.lastname: Foo
authors.affiliation: Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore 119260, Singapore.
authors.forename: J
authors.initials: J
authors.lastname: Sivaraman
authors.affiliation: Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore 119260, Singapore.
authors.forename: R Manjunatha
authors.initials: RM
authors.lastname: Kini
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000925:Anticoagulants / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology / Q000633:toxicity; D002645:Chickens; D018360:Crystallography, X-Ray; D003546:Cysteine Endopeptidases; D005170:Factor X; D000073179:Hemachatus / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D051379:Mice; D008958:Models, Molecular; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors*; D010264:Paraplegia / Q000139:chemically induced*; D017433:Protein Structure, Secondary; D012910:Snake Venoms / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology / Q000633:toxicity
chemical_list: D000925:Anticoagulants; D009363:Neoplasm Proteins; D012910:Snake Venoms; D005170:Factor X; D003546:Cysteine Endopeptidases; C030910:cancer procoagulant