title: A frequent human coagulation Factor VII mutation (A294V, c152) in loop 140s affects the interaction with activators, tissue factor and substrates.
abstract: Activated Factor VII (FVIIa) is a vitamin-K-dependent serine protease that initiates blood clotting after interacting with its cofactor tissue factor (TF). The complex FVIIa-TF is responsible for the activation of Factor IX (FIX) and Factor X (FX), leading ultimately to the formation of a stable fibrin clot. Activated FX (FXa), a product of FVIIa enzymic activity, is also the most efficient activator of zymogen FVII. Interactions of FVII/FVIIa with its activators, cofactor and substrates have been investigated extensively to define contact regions and residues involved in the formation of the complexes. Site-directed mutagenesis and inhibition assays led to the identification of sites removed from the FVIIa active site that influence binding specificity and affinity of the enzyme. In this study we report the characterization of a frequent naturally occurring human FVII mutant, A294V (residue 152 in the chymotrypsin numbering system), located in loop 140s. This region undergoes major rearrangements after FVII activation and is relevant to the development of substrate specificity. FVII A294V shows delayed activation by FXa as well as reduced activity towards peptidyl and macromolecular substrates without impairing the catalytic efficiency of the triad. Also, the interaction of this FVII variant with TF was altered, suggesting that this residue, and more likely loop 140s, plays a pivotal role not only in the recognition of FX by the FVIIa-TF complex, but also in the interaction of FVII with both its activators and cofactor TF.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Ferrara, Via Borsari, 46 Ferrara 44100, Italy. r_toso@hotmail.com
authors.forename: Raffaella
authors.initials: R
authors.lastname: Toso
authors.forename: Mirko
authors.initials: M
authors.lastname: Pinotti
authors.forename: Katherine A
authors.initials: KA
authors.lastname: High
authors.forename: Eleanor S
authors.initials: ES
authors.lastname: Pollak
authors.forename: Francesco
authors.initials: F
authors.lastname: Bernardi
mesh_terms: D001665:Binding Sites; D001777:Blood Coagulation; D010450:Endopeptidases / Q000378:metabolism; D005167:Factor VII / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D015942:Factor VIIa / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014644:Genetic Variation; D006801:Humans; D066298:In Vitro Techniques; D017354:Point Mutation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D013925:Thromboplastin / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D005167:Factor VII; D013925:Thromboplastin; D010450:Endopeptidases; D015942:Factor VIIa