Coagulation Factor V
Coagulation factor V (FV) circulates in the bloodstream in an inactive form (procofactor) and is activated to factor Va (FVa) by thrombin. Thrombin cleaves away the large inhibitory B-domain of FV which resolves the molecule into a heterodimer that is stabilized through non-covalent interactions between heavy (A1-A2) and light (A3-C1-C2) chains. A recombinantly expressed truncated B-domain variant of factor V (FV-DT) exhibits constitutive FVa-like activity even in the absence of proteolysis. FV maintains its cofactor state through a bipartite autoinhibitory system present in B-domain. Biochemical evidence indicates that a basic fragment (BR) of B-domain interacts with at least one of the two acidic regions present at the two flanks of B-domain
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The two C-domains are oriented just as seen in bFVai structure, however they differ in crystal contacts with the neighboring molecules in the crystal. The structure shows the presence of two calcium ions, one in A1 and the other in A3 domain. The absence of Ca2+ in A3 domain in bFVai structure indicates a structural destabilization of the Ca-binding loop that accompanies the release of A2 domain upon inactivation of FVa by APC. The most important aspect of the structure is a close spatial proximity of the two conserved acidic regions that are bisected by the long intervening B-domain in the primary sequence. Our observations provide the first structural evidence that the two distinct acidic regions come together in space to provide an extended surface to which the basic fragment may bind to maintain FV in a procofactor state. Interestingly, this acidic surface lies at the interface of the FVa and Xa interaction, which explains why occlusion of this site by BR or TFPIα can efficiently interfere with the prothrombinase assembly. Furthermore, the calcium-binding loop in the A3-domain lies close to the bisegmental acidic cluster present at the junction of A2 and A3 domain. Consequently, any small perturbation in the Ca-binding loop in the A3-domain may have severe implications on the protein's functionality. The loops containing two APC cleavage sites in the A2 domain (Arg 306 and Arg506) were not completely modeled in the structure, suggesting their high flexibility may be required for them to be readily accessible to the active site of
The two C-domains are oriented just as seen in bFVai structure, however they differ in crystal contacts with the neighboring molecules in the crystal. The structure shows the presence of two calcium ions, one in A1 and the other in A3 domain. The absence of Ca2+ in A3 domain in bFVai structure indicates a structural destabilization of the Ca-binding loop that accompanies the release of A2 domain upon inactivation of FVa by APC. The most important aspect of the structure is a close spatial proximity of the two conserved acidic regions that are bisected by the long intervening B-domain in the primary sequence. Our observations provide the first structural evidence that the two distinct acidic regions come together in space to provide an extended surface to which the basic fragment may bind to maintain FV in a procofactor state. Interestingly, this acidic surface lies at the interface of the FVa and Xa interaction, which explains why occlusion of this site by BR or TFPIα can efficiently interfere with the prothrombinase assembly. Furthermore, the calcium-binding loop in the A3-domain lies close to the bisegmental acidic cluster present at the junction of A2 and A3 domain. Consequently, any small perturbation in the Ca-binding loop in the A3-domain may have severe implications on the protein's functionality. The loops containing two APC cleavage sites in the A2 domain (Arg 306 and Arg506) were not completely modeled in the structure, suggesting their high flexibility may be required for them to be readily accessible to the active site of