ABSTRACT
Human coagulation factor V (FV), a non-enzymatic cofactor of the prothrombinase complex, is required for the rapid generation of thrombin. FV deficiency is a rare autosomal recessive bleeding disorder. We describe two novel mutations, Tyr91Asn and Asp2098Tyr, found in two probands with a residual FV activity of 51% and 4%, respectively. Modelling and structural analysis of these mutations were performed following short-duration molecular dynamics (MD) simulation. Asp2098Tyr lead to abolishment of the highly conserved salt bridge Asp2098-Arg2171 presumably required for structural integrity of the C2 domain. MD studies suggest that additional conformational changes resulting from this mutation involve local rearrangements at Tyr2063 and Tyr2064 and so affect the phospholipid-membrane binding. MD modelling of the Try91Asn mutant revealed a conformational change nearby the Cu(2+) binding site that could affect overall stabilization of the heavy and light chains. These findings suggest that both mutations influence the structural integrity of FV protein. Transient expression data of wild-type and mutant FV variants in 293T human embryonic kidney cells showed FV-specific activity reduced to 26% for Asp2098Tyr and 56% for Tyr91Asn compared to that of wild-type. Thus, both the data from the short duration molecular dynamic simulation and from expression analysis indicate alterations of the FV protein variants that explain the clinical phenotype.
