Steroid 17alpha-hydroxylase deficiency: functional characterization of four mutations (A174E, V178D, R440C, L465P) in the CYP17A1 gene

CONTEXT: Steroid 17alpha-hydroxylase (CYP17A1, alias P450c17) deficiency (17OHD) is a rare form of congenital adrenal hyperplasia. The CYP17A1 enzyme catalyzes two distinct reactions, 17alpha-hydroxylase and 17,20-lyase activities. OBJECTIVE: The aim of the study was to analyze the structural and functional consequences of three novel (A174E, V178D, and L465P) and one previously reported (R440C) CYP17A1 mutation found in three patients clinically and biochemically presenting with 17OHD. PATIENTS AND METHODS: Two patients suffering from 46,XY disordered sex development presented at ages 5.5 and 8.8 yr, respectively, with tall stature and hypertension. Mutation analysis revealed compound heterozygous CYP17A1 mutations (A174E/K388X; V178D/R440C). The third patient (46,XX) presented with primary amenorrhea and hypertension at age 15 yr. She was homozygous for the novel L465P mutation. Functional studies employing a yeast microsomal expression system compared wild-type and mutant CYP17A1 both with regard to 17alpha-hydroxylase and 17,20-lyase activity. Mutants were examined in a computational three-dimensional model of the CYP17A1 protein. RESULTS: The activity assays showed that all three mutants retain only 0-7% of both 17alpha-hydroxylase and 17,20-lyase activity relative to CYP17A1 wild-type activity, corresponding to the in vivo situation. Enzyme kinetic studies proved the impairment of both reactions, respectively. Computer-based three-dimensional model analysis of CYP17A1 using CYP2B4 as template showed that three of the mutations had no direct effect on the active center, whereas one affects the heme coordination. CONCLUSION: The functional studies revealed that the described missense mutations result in severe 17OHD. Our data are important to predict the phenotypic expressions and provide important information for patient management and genetic counseling.