Brenner B, Sánchez-Vega B, Wu S M, Lanir N, Stafford D W, Solera J
Thrombosis and Hemostasis Unit, Institute of Hematology, Rambam Medical Center, and Bruce Rappaport Faculty of Medicine, Technion, Haifa, Israel.
Blood. 1998 Dec 15;92(12):4554-9.
To identify potential mutations in the gamma-glutamyl carboxylase gene, the sequence of all exons and intron/exon borders was determined in 4 patients from a consanguineous kindred with combined deficiency of all vitamin K-dependent procoagulants and anticoagulants and results were compared with normal genomic sequence. All 4 patients were homozygous for a point mutation in exon 9 that resulted in the conversion of an arginine codon (CTG) to leucine codon (CGG) at residue 394. Screening of this mutation based on introduction of Alu I site in amplified fragment from normal allele but not from the mutated allele showed that 13 asymptomatic members of the kindred were heterozygous for the mutation. The mutation was not found in 340 unrelated normal chromosomes. The segregation pattern of the mutation which is the first reported in the gamma-glutamyl carboxylase gene fits perfectly with phenotype of the disorder and confirms the suggested autosomal recessive pattern of inheritance of combined deficiency of all vitamin K-dependent procoagulants and anticoagulants in this kindred. The mutated carboxylase protein expressed in Drosophila cells was stable but demonstrated threefold reduced activity compared with WT carboxylase, confirming that the L394R mutation results in a defective carboxylase.
为了鉴定γ-谷氨酰羧化酶基因中的潜在突变，对来自一个近亲家族的4名患者的所有外显子以及内含子/外显子边界的序列进行了测定，这些患者存在所有维生素K依赖的促凝血因子和抗凝血因子联合缺乏的情况，并将结果与正常基因组序列进行了比较。所有4名患者在外显子9中均为一个点突变的纯合子，该突变导致第394位残基处的精氨酸密码子（CTG）转变为亮氨酸密码子（CGG）。基于在正常等位基因而非突变等位基因的扩增片段中引入Alu I位点对该突变进行筛查，结果显示该家族的13名无症状成员为该突变的杂合子。在340条无关的正常染色体中未发现该突变。γ-谷氨酰羧化酶基因中首次报道的这种突变的分离模式与该疾病的表型完全相符，并证实了该家族中所有维生素K依赖的促凝血因子和抗凝血因子联合缺乏所提示的常染色体隐性遗传模式。在果蝇细胞中表达的突变型羧化酶蛋白是稳定的，但与野生型羧化酶相比活性降低了三倍，这证实了L394R突变导致羧化酶缺陷。
