title: The roles of factor VII's structural domains in tissue factor binding.
abstract: Factor VIIa binds to tissue factor in one of the initial steps of blood clotting. In order to determine the role of the various domains of the factor VII molecule in this interaction, we made several chimeric factor VII proteins using recombinant DNA techniques. The molecules have factor IX domains substituted into factor VII and vice versa. The domains exchanged were the 4-carboxyglutamic acid plus aromatic stack domain (gla), the first epidermal growth factor-like domain (Egf-1), the second epidermal growth factor-like domain (Egf-2), and the catalytic domain. Using tissue factor-coated microtiter wells, competition binding studies with 125I-labeled factor VIIa indicated factor VIIa's Kd is 4.2 nM. Employing the same microtiter plate assay, koff and kon were determined and yielded a Kd of 1.5 nM. The results of competitive binding experiments and activation assays using chimeric proteins indicated the interaction between factor VIIa and tissue factor involves direct contact between tissue factor and factor VIIa's Egf-1 domain and catalytic domain. On the other hand, the gla and Egf-2 domains, while necessary for optimal binding, may merely impart structure to the rest of the molecule. However, either one or both of the latter domains might contribute a relatively small amount of energy to direct binding.
authors.affiliation: Department of Biology, University of North Carolina at Chapel Hill 27599, USA.
authors.forename: J Y
authors.initials: JY
authors.lastname: Chang
authors.forename: D W
authors.initials: DW
authors.lastname: Stafford
authors.forename: D L
authors.initials: DL
authors.lastname: Straight
mesh_terms: D001667:Binding, Competitive; D005164:Factor IX / Q000235:genetics / Q000378:metabolism; D015942:Factor VIIa / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D011485:Protein Binding; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship; D013925:Thromboplastin / Q000378:metabolism*
chemical_list: D011993:Recombinant Fusion Proteins; D005164:Factor IX; D013925:Thromboplastin; D015942:Factor VIIa