title: Coagulation factor X-binding protein from Deinagkistrodon acutus venom is a Gla domain-binding protein.
abstract: Factor IX/factor X-binding protein (IX/X-bp) is an anticoagulant isolated from the venom of Trimeresurus flavoviridis (habu snake) and binds predominantly to factor IX. In this study, we isolated IX/X-bp-like proteins from the venom of Deinagkistrodon acutus (hundred pace snake) with binding characteristics different from those of IX/X-bp. The complete amino acid sequence and binding characteristics of the main anticoagulant protein, named X-bp, were investigated. The concentrations of X-bp at half-maximal binding to solid-phase factors X and IX were 0.4 and 3 nM, respectively. The binding of X-bp to solid-phase factor X was inhibited by 50% by 6- and 9-fold excess concentrations of factor X and Gla domain (GD) peptide 1-44, respectively, but was not influenced by GD peptide 1-41 and Gla domainless factor X. X-bp bound two Ca2+ ions per molecule with Kd values of 16 +/- 0.7 (mean +/- SE, n = 6) and 103 +/- 10 microM. X-bp was a heterodimer of C-type lectin-like subunits. The 16 kDa chain (A chain) consisted of 129 amino acid residues and was 68% identical to the sequence of the A chain of IX/X-bp. The 15 kDa chain (B chain) consisted of 123 amino acid residues and was 87% identical to IX/X-bp. Three-dimensional model construction from the known fold of IX/X-bp showed that amino acid residues different from those of IX/X-bp are mostly on the molecular surface. Some of these are concentrated on a part of the concave surface which is considered to be the coagulation factor-binding site, presumably acting as a discriminator for ligand binding. These results indicated that X-bp isolated from D. acutus venom was a GD-binding protein, and the C-terminal region of GD peptide was critical for folding of the peptide.
authors.affiliation: Department of Biochemistry, Meiji Pharmaceutical University, Tokyo, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Atoda
authors.forename: M
authors.initials: M
authors.lastname: Ishikawa
authors.forename: H
authors.initials: H
authors.lastname: Mizuno
authors.forename: T
authors.initials: T
authors.lastname: Morita
mesh_terms: D015055:1-Carboxyglutamic Acid / Q000378:metabolism*; D017836:Agkistrodon; D000595:Amino Acid Sequence; D000818:Animals; D000925:Anticoagulants / Q000737:chemistry / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002417:Cattle; D003435:Crotalid Venoms / Q000737:chemistry / Q000378:metabolism*; D005170:Factor X / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D011485:Protein Binding; D030162:Reptilian Proteins; D017421:Sequence Analysis
chemical_list: D000925:Anticoagulants; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D003435:Crotalid Venoms; D010446:Peptide Fragments; D030162:Reptilian Proteins; C070302:factor IX-factor X-binding protein, Crotalinae; D015055:1-Carboxyglutamic Acid; D005170:Factor X; D002118:Calcium