Riepe Felix G, Hiort Olaf, Grötzinger Joachim, Sippell Wolfgang G, Krone Nils, Holterhus Paul-Martin
Division of Pediatric Endocrinology, Department of Pediatrics, University Hospital Schleswig-Holstein, Schwanenweg 20, Kiel, Germany.
J Clin Endocrinol Metab. 2008 Jul;93(7):2891-5. doi: 10.1210/jc.2007-2646. Epub 2008 Apr 29.
Congenital adrenal hyperplasia is caused by insufficient adrenal steroid biosynthesis due to impaired steroidogenic enzymes. The majority of patients suffer from deficiency of 21-hydroxylase (CYP21) coded by the CYP21A2 gene.
Our objective was to study the functional and structural consequences of the novel CYP21A2 missense mutation c.364A > C (K121Q) detected in a female patient with nonclassical 21-hydroxylase deficiency. The patient was compound heterozygous for the novel K121Q mutation and the mild P453S mutation.
In vitro expression analysis of the mutant K121Q enzyme in transiently transfected COS-7 cells revealed reduced CYP21 activity of 14.0 +/- 5% for the conversion of 17-hydroxyprogesterone and 19.5 +/- 4% for the conversion of progesterone. K121 is located on helix C in the CYP21 protein, which is part of the heme coordinating system. In addition, helix C is involved in the interaction with the electron-providing enzyme P450 oxidoreductase. Protein modeling revealed that the substitution of glutamine for the basic amino acid lysine introduces an electrostatic change on the surface of CYP21 and may additionally change heme coordination. We hypothesize that the electron flux between P450 oxidoreductase and CYP21 is impaired and, moreover, that substrate affinity is altered due to heme dislocation with K121Q.
Both the interaction of P450 oxidoreductase and CYP21 as well as heme coordination are likely to be disturbed due to the K121Q mutation. Our data exemplify how the combination of in vitro expression and structural protein analysis provide novel insights into molecular mechanisms of reduced CYP21 activity, eventually explaining the patient's phenotype.
先天性肾上腺皮质增生症是由于类固醇生成酶受损导致肾上腺类固醇生物合成不足所致。大多数患者患有由CYP21A2基因编码的21-羟化酶（CYP21）缺乏症。
我们的目的是研究在一名非经典21-羟化酶缺乏症女性患者中检测到的新型CYP21A2错义突变c.364A>C（K121Q）的功能和结构后果。该患者为新型K121Q突变和轻度P453S突变的复合杂合子。
在瞬时转染的COS-7细胞中对突变型K121Q酶进行体外表达分析，结果显示17-羟孕酮转化的CYP21活性降低了14.0±5%，孕酮转化的活性降低了19.5±4%。K121位于CYP21蛋白的C螺旋上，C螺旋是血红素配位系统的一部分。此外，C螺旋参与与提供电子的酶P450氧化还原酶的相互作用。蛋白质建模显示，用谷氨酰胺取代碱性氨基酸赖氨酸会在CYP21表面引入静电变化，并且可能还会改变血红素配位。我们推测P450氧化还原酶和CYP21之间的电子通量受损，此外，由于K121Q导致血红素错位，底物亲和力也发生了改变。
由于K121Q突变，P450氧化还原酶和CYP21之间的相互作用以及血红素配位可能都会受到干扰。我们的数据例证了体外表达和结构蛋白分析相结合如何为CYP21活性降低的分子机制提供新的见解，最终解释了患者的表型。
