17α-羟化酶缺乏症的分子遗传学研究

Molecular genetic study on 17α-hydroxylase deficiency

目的 研究一例 17α 羟化酶 (CYP17)缺乏症患者发病的分子遗传学机制。方法 从全血中抽提患者及其父母亲和正常人基因组DNA并用预先设计的 5对引物通过多聚酶链反应(PCR)扩增CYP17基因的 8个外显子 ,用双脱氧链终止法测定患者CYP17基因 8个外显子序列 ,并进行同源性比较 ,以确定其突变位点 ;测定患者父、母亲的相应序列 ,确定突变的遗传来源。结果 患者CYP17基因存在复合的杂合突变 :一个突变位点来自母亲 ,即外显子 1中Arg96 (CGG)→Gln(CAG) ;另一个突变位点来自父亲 ,系外显子 8的第 487～ 489位密码子GACTCTTTC共 9个碱基缺失 ,导致编码该处的Asp4 87 Ser4 88 Phe4 89共 3个氨基酸残基缺失。结论 该患者 17α 羟化酶活性缺陷的原因极可能是由于CYP17基因中存在上述复合的杂合突变 ,导致P45 0c17酶氨基酸序列改变而影响酶活性。P45 0c17酶蛋白一级结构中第 96位精氨酸对维持P45 0c17酶活性至关重要。外显子 8中Asp4 87 Ser4 88 Phe4

Objective To study the molecular genetic mechanism of a patient with 17α hydroxylase (CYP17) deficiency. Methods Genomic DNA were abstracted from the blood of the patient, her parents and healthy control. The 8 exons of CYP17 gene were amplified, using 5 pairs of designed primers, with polymerase chain reaction (PCR), and the 8 exons were sequenced by the dideoxy terminator method to determined the mutation sites. The corresponding exons of the parents of the patients were also amplified and sequenced to determine the zygosity of the patient and the source of the gene variances. Results The analysis revealed that the patient (46, XY) was a compound heterozygote carrying two different inherited mutations on CYP17 gene, one from mother containing a point mutation Arg 96 (C G G)→ Gln(C A G) and the other from father containing a nine base deletion (CACTCTTTC) at amino acid position 487～489 (Asp Ser Phe) near the carboxyl terminus of P450c17. Conclusion The CYP17 gene of the patient with 17α hydroxylase deficiency is a compound heterozygous mutation. The mutation changes the amino acid sequence of P450c17 enzyme, which in turn affected the enzymatic activity. Arg 96 is essential in P450c17 enzyme activity. Deletion of Asp 487 Ser 488 Phe 489 in exon 8 may be a prevalent mutation causing P450c17 deficiency in Southeast Asia.