Sun Bang, Lu Lin, Xie Shaowei, Zhang Wei, Zhang Xiaoxia, Tong Anli, Chen Shi, Wu Xueyan, Mao Jiangfeng, Wang Xi, Qiu Ling, Nie Min
Department of Endocrinology, NHC Key laboratory of Endocrinology (Peking Union Medical College Hospital), Peking Union Medical College Hospital, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China.
Department of Clinical Laboratory, Peking Union Medical College Hospital, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China.
FASEB J. 2023 Apr;37(4):e22869. doi: 10.1096/fj.202201398RR.
Steroid 11β-hydroxylase deficiency (11β-OHD) is a rare autosomal recessive disorder caused by pathogenic variants of CYP11B1 gene. This study aimed to perform molecular analysis of a Chinese 11β-OHD series and in vitro functional study of twenty CYP11B1 missense variants. Twelve Chinese patients with clinical diagnosis of 11β-OHD were included in the study to analyze their molecular etiology. Genomic DNA of patients was extracted to be sequenced all coding exons and intronic flanking sequences of CYP11B1. Fourteen missense variants found in 12 patients mentioned above along with 6 missense variants previously reported by our team were evaluated functionally. Amino acid substitutions were analyzed with computational program to determine their effects on the three-dimensional structure of CYP11B1 protein. Clinical characteristics and hormone levels at baseline of the 18 patients carrying 18 missense variants aforementioned were recorded to perform genotype-phenotype correlation. A total of 21 rare variants including 9 novel and 12 recurrent ones were identified in 12 patients, out of which 17 were missense, 2 were nonsense, 1 was a splice site variant, and 1 was a deletion-insertion variant. Results of in vitro functional study revealed that 3 out of 20 missense mutants (p.Leu3Pro, p.Gly267Ser, and p.Ala367Ser) had partial enzyme activity and the other 17 had little enzymatic activity. The impairment degree of enzymatic activity in vitro functional study was also reflected in the severity degree of interaction change between the wild-type/mutant-type amino acid and its adjacent amino acids in three-dimensional model. In conclusion, the addition of 9 novel variants expands the spectrum of CYP11B1 pathogenic variants. Our results demonstrate that twenty CYP11B1 variants lead to impaired 11β-hydroxylase activity in vitro. Visualizing these variants in the three-dimensional model structure of CYP11B1 protein can provide a plausible explanation for the results measured in vitro.
类固醇11β-羟化酶缺乏症（11β-OHD）是一种由CYP11B1基因的致病变异引起的罕见常染色体隐性疾病。本研究旨在对一组中国11β-OHD患者进行分子分析，并对20个CYP11B1错义变体进行体外功能研究。本研究纳入了12例临床诊断为11β-OHD的中国患者，以分析其分子病因。提取患者的基因组DNA，对CYP11B1的所有编码外显子和内含子侧翼序列进行测序。对上述12例患者中发现的14个错义变体以及我们团队先前报道的6个错义变体进行功能评估。使用计算程序分析氨基酸取代，以确定它们对CYP11B1蛋白三维结构的影响。记录携带上述18个错义变体的18例患者的临床特征和基线激素水平，以进行基因型-表型相关性分析。在12例患者中总共鉴定出21个罕见变体，包括9个新变体和12个复发变体，其中17个是错义变体，2个是无义变体，1个是剪接位点变体，1个是缺失-插入变体。体外功能研究结果显示，20个错义突变体中有3个（p.Leu3Pro、p.Gly267Ser和p.Ala367Ser）具有部分酶活性，其他17个几乎没有酶活性。体外功能研究中酶活性的受损程度也反映在三维模型中野生型/突变型氨基酸与其相邻氨基酸之间相互作用变化的严重程度上。总之，9个新变体的加入扩大了CYP11B1致病变异的谱。我们的结果表明，20个CYP11B1变体在体外导致11β-羟化酶活性受损。在CYP11B1蛋白的三维模型结构中可视化这些变体可以为体外测量结果提供合理的解释。
