[Phenotypic and mutational analysis of a pedigree affected with hereditary coagulation factor Ⅴ deficiency]

To explore the molecular pathogenesis for a pedigree affected with coagulation factor Ⅴ (FⅤ) deficiency.

Prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB), coagulation factor Ⅱ activity (FⅡ: C), FⅤ activity (FⅤ: C), coagulation factor Ⅶ activity (FⅦ: C), and coagulation factor Ⅹ activity (FⅩ: C) were determined with a STAGO automatic coagulometer. FⅤ antigen (FⅤ: Ag) was detected with enzyme linked immunosorbent assay (ELISA). All exons and their flanking regions, and 5' and 3' untranslated regions of the F5 gene were analyzed by direct sequencing. Suspected mutation was verified by reverse sequencing as well as testing of family members. ClustalX software was used to analyze the conservative property of the mutation sites. PROVEAN and MutationTaster online software was used to predict the effect of the mutation on the protein function. Swiss-pdbViewer was used to analyze the protein model and interaction of amino acids.

The PT and APTT of the proband were slightly prolonged to 15.2 s and 41.8 s, respectively. And the FⅤ: C and FⅤ: Ag measured 55% and 62%, respectively. The FⅤ: C and FⅤ: Ag of his father and son were decreased to various extent (60%, 65% and 31%, 40%, respectively). A c.911G>A heterozygous mutation (Gly276Glu) was detected in exon 6 of the proband, for which her father and son were heterozygotes. The same mutation was not found in her mother, brother and husband. Conservation analysis showed that the Gly276 is highly conserved across various species. By bioinformatic analysis, the PROVEAN (scored -6.214) indicated Gly276Glu was harmful, and MutationTaster (scored 0.976) suggested that it is pathogenic. Model analysis suggested there are two hydrogen bonds between Gly276 and Ile298 in the wild type protein. When Gly276 was replaced by Glu276, the original hydrogen bond did not change, but the side chain of Glu was extended, which added steric hindrance with the surrounding amino acids, which resulted in decreased protein stability.

The heterozygous c.911G>A (Gly276Glu) mutation of the F5 gene probably underlies the decreased level of FⅤin the proband.