FACTORS X(WENATCHEE)-I AND X(WENATCHEE)-II - COMPOUND HETEROZYGOSITY INVOLVING 2 VARIANT PROTEINS

Variant factor X in an individual with a mild bleeding tendency was su spected based on deficient procoagulant activity (10-20% of normal) an d antigen (30-35% of normal) levels of plasma factor X. Heteroduplex a nalysis of factor X gene exons indicated heterozygosity for mutations in both exons 6 and 4, confirmed by direct sequencing of the amplified exons. Substitution of C by T at nucleotide position 13984 (Arg-139 t o Cys) was found in the factor X gene exon 6 of the propositus. This m utation creates a BsmI site and the patient tested heterozygous for th e BsmI cleavage involved, as did one of his two daughters. In addition , exon 4 was found to have the normal A and a novel C (Asn-57 to Thr) at nucleotide position 9338. The exon 4 mutation creates a BsaJI site, detectable after amplification mismatch to remove an existing BsaJI s ite. Both the patient and the second of his two daughters were heteroz ygous for this cleavage. The two variant proteins are called factors X (Wenatchee) I (Arg-139 to Cys) and II (Asn-57 to Thr). A mixed variant isolate derived from the plasma of the propositus exhibited heavy/lig ht chains of normal size, as well as an apparent single-chain molecule not dissociable by reducing agent. A single-chain molecule would be p redicted for form I, if the mutation blocks processing cleavages that normally remove a tripeptide interposed between the heavy and light ch ains. A Western blot of partially purified factor X from the daughter who inherited the form I defect revealed a component migrating the sam e as the putative single-chain species. Based upon the factor X activi ty vs. antigen ratios for the propositus and both daughters, both form s I and II are probably dysfunctional molecules.