1 Université Paris-Sud , Orsay, France .
2 UMR 8200 CNRS , Villejuif, France .
Thyroid. 2018 Aug;28(8):1052-1062. doi: 10.1089/thy.2017.0596. Epub 2018 Jul 24.
Dual oxidases (DUOX1 and DUOX2) were initially identified as HO sources involved in thyroid hormone synthesis. Congenital hypothyroidism (CH) resulting from inactivating mutations in the DUOX2 gene highlighted that DUOX2 is the major HO provider to thyroperoxidase. The role of DUOX1 in the thyroid remains unknown. A recent study suggests that it could compensate for DUOX2 deficiency in CH. Both DUOX enzymes and their respective maturation factors DUOXA1 and DUOXA2 form a stable complex at the cell surface, which is fundamental for their enzymatic activity. Recently, intra- and intermolecular disulfide bridges were identified that are essential for the structure and the function of the DUOX2-DUOXA2 complex. This study investigated the involvement of cysteine residues conserved in DUOX1 toward the formation of disulfide bridges, which could be important for the function of the DUOX1DUOXA1 complex.
To analyze the role of these cysteine residues in both the targeting and function of dual oxidase, different human DUOX1 mutants were constructed, where the cysteine residues were replaced with glycine. The effect of these mutations on cell surface expression and HO-generating activity of the DUOX1-DUOXA1 complex was analyzed.
Mutations of two cysteine residues (C118 and C1165), involved in the formation of the intramolecular disulfide bridge between the N-terminal ectodomain and one of the extracellular loops, mildly altered the function and the targeting of DUOX1, while this bridge is crucial for DUOX2 function. Unlike DUOXA2, with respect to DUOX2, the stability of the maturation factor DUOXA1 is not dependent on the oxidative folding of DUOX1. Only mutation of C579 induced a strong alteration of both targeting and function of the oxidase by preventing the covalent interaction between DUOX1 and DUOXA1.
An intermolecular disulfide bridge rather than an intramolecular disulfide bridge is important for both the trafficking and HO-generating activity of the DUOX1-DUOXA1 complex.
双氧化酶（DUOX1 和 DUOX2）最初被鉴定为参与甲状腺激素合成的 HO 来源。DUOX2 基因失活突变导致的先天性甲状腺功能减退症（CH）突出表明 DUOX2 是甲状腺过氧化物酶的主要 HO 供体。DUOX1 在甲状腺中的作用尚不清楚。最近的一项研究表明，它可以弥补 CH 中 DUOX2 缺乏。DUOX 酶及其各自的成熟因子 DUOXA1 和 DUOXA2 在细胞表面形成稳定的复合物，这对于它们的酶活性至关重要。最近，鉴定了二硫键在内和分子间的二硫键，这对于 DUOX2-DUOXA2 复合物的结构和功能是必不可少的。本研究调查了 DUOX1 中的半胱氨酸残基对形成二硫键的参与情况，这对于 DUOX1DUOXA1 复合物的功能可能很重要。
为了分析这些半胱氨酸残基在双氧化酶的靶向和功能中的作用，构建了不同的人 DUOX1 突变体，其中半胱氨酸残基被甘氨酸取代。分析这些突变对 DUOX1-DUOXA1 复合物的细胞表面表达和 HO 生成活性的影响。
两个半胱氨酸残基（C118 和 C1165）的突变，参与了 N 端胞外域和其中一个细胞外环之间的分子内二硫键的形成，轻度改变了 DUOX1 的功能和靶向，而该桥对于 DUOX2 功能至关重要。与 DUOXA2 不同，就 DUOX2 而言，成熟因子 DUOXA1 的稳定性不依赖于 DUOX1 的氧化折叠。只有 C579 的突变通过阻止 DUOX1 和 DUOXA1 之间的共价相互作用，强烈改变了氧化酶的靶向和功能。
对于 DUOX1-DUOXA1 复合物的转运和 HO 生成活性，分子间二硫键比分子内二硫键更为重要。
