Factor VIIa: Difference between revisions
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The binding epitope of TF to FVIIa is a stripe running along the whole length of the TF protein. | The binding epitope of TF to FVIIa is a stripe running along the whole length of the TF protein. | ||
The Gla domain of FVIIa binds to the C-domain of TF. The interaction is mainly hydrophobic termed the “hydrophobic stack”. EGF1 binds a single Ca++ ion and packs into a groove formed by the two modules of TF. This interface is the largest and contributes the most energetically in binding of the cofactor to the protease domain13. EGF2 and the catalytic domain interact with the N-domain of TF. Two “lock and key” interactions are observed. One is the side chain of Phe50 of TF which is trapped in the pocket formed by the end of EGF2 domain of FVIIa. Second, the side chain Met306 of FVIIa is enclosed by TF residues Arg74, Phe76, Glu92, Leu94. This Met306 residue is responsible for the thermodynamic coupling to the active site and is unique to FVIIa. Mutation of this residue results in the failure of TF to decrease the dissociation rate of the enzyme from the cofactor. Cofactor interactions, specifically through Met306 lead to subtle changes which then influence the position of Asp331 (check). Asp331 is the specificity-determining residue in the binding pocket. Constraint and stabilization promote formation of a hydrogen bond between the amide of Arg315 (170C) and carbonyl of Gly372 (223). When bound to TF the activation region of FVIIa contains a large number of hydrogen bonds between the main chain and side chain atoms. At least 11 water molecules are identified in the catalytic domain region. An interesting observation is that the center of this region contains water, in contrast to the hydrophobic interactions in the other two interface regions. These hydrophilic interactions may be more efficient at mediating TF affinity and substantial conformational changes to induce activity of FVIIa. Therefore TF and cations are obligatory cofactors in the allosteric regulation of FVIIa activity by stabilizing the disordered, felxible FVIIa and restraining the enzyme for catalytic activity. | The Gla domain of FVIIa binds to the C-domain of TF. The interaction is mainly hydrophobic termed the “hydrophobic stack”. EGF1 binds a single Ca++ ion and packs into a groove formed by the two modules of TF. This interface is the largest and contributes the most energetically in binding of the cofactor to the protease domain13. EGF2 and the catalytic domain interact with the N-domain of TF. Two “lock and key” interactions are observed. One is the side chain of Phe50 of TF which is trapped in the pocket formed by the end of EGF2 domain of FVIIa. Second, the side chain Met306 of FVIIa is enclosed by TF residues Arg74, Phe76, Glu92, Leu94. This Met306 residue is responsible for the thermodynamic coupling to the active site and is unique to FVIIa. Mutation of this residue results in the failure of TF to decrease the dissociation rate of the enzyme from the cofactor. Cofactor interactions, specifically through Met306 lead to subtle changes which then influence the position of Asp331 (check). Asp331 is the specificity-determining residue in the binding pocket. Constraint and stabilization promote formation of a hydrogen bond between the amide of Arg315 (170C) and carbonyl of Gly372 (223). When bound to TF the activation region of FVIIa contains a large number of hydrogen bonds between the main chain and side chain atoms. At least 11 water molecules are identified in the catalytic domain region. An interesting observation is that the center of this region contains water, in contrast to the hydrophobic interactions in the other two interface regions. These hydrophilic interactions may be more efficient at mediating TF affinity and substantial conformational changes to induce activity of FVIIa. Therefore TF and cations are obligatory cofactors in the allosteric regulation of FVIIa activity by stabilizing the disordered, felxible FVIIa and restraining the enzyme for catalytic activity. | ||
===Kinetics of action=== | |||
In general serine proteases have a three-step kinetic mechanim: 1) formation of an enzyme-substrate (E-S) complex, 2) acylation of the active site serine, and 3) hydrolysis of the acylenzyme intermediate. A large solvent isotope effect (~3) | |||
[[Image:kinetics ser protease.jpg|center]] | |||
===Catalytic domain=== | ===Catalytic domain=== | ||
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Upon attack on an amide bond, the lone electron pairs of the oxyanion and the nitrogen of the leaving group have to be antiperiplanar to the new bond. Therefore the lone pair of the amine leaving group points away from His193-H+. For the chemistry to occur the nitrogen must undergo inversion to position the lone pair for protonation by His193-H+<ref>PMID:2514538</ref>. | Upon attack on an amide bond, the lone electron pairs of the oxyanion and the nitrogen of the leaving group have to be antiperiplanar to the new bond. Therefore the lone pair of the amine leaving group points away from His193-H+. For the chemistry to occur the nitrogen must undergo inversion to position the lone pair for protonation by His193-H+<ref>PMID:2514538</ref>. | ||
[[Image:stereochemistry of N.jpg|center]] | [[Image:stereochemistry of N.jpg|center]] | ||
==References== | ==References== | ||
<references /> | <references /> | ||