Department of Biochemistry, Medical College of Wisconsin, WI, USA.
Arch Biochem Biophys. 2012 Dec 1;528(1):72-89. doi: 10.1016/j.abb.2012.09.002. Epub 2012 Sep 11.
NADPH-cytochrome P450 oxidoreductase (CYPOR) and nitric oxide synthase (NOS), two members of the diflavin oxidoreductase family, are multi-domain enzymes containing distinct FAD and FMN domains connected by a flexible hinge. FAD accepts a hydride ion from NADPH, and reduced FAD donates electrons to FMN, which in turn transfers electrons to the heme center of cytochrome P450 or NOS oxygenase domain. Structural analysis of CYPOR, the prototype of this enzyme family, has revealed the exact nature of the domain arrangement and the role of residues involved in cofactor binding. Recent structural and biophysical studies of CYPOR have shown that the two flavin domains undergo large domain movements during catalysis. NOS isoforms contain additional regulatory elements within the reductase domain that control electron transfer through Ca(2+)-dependent calmodulin (CaM) binding. The recent crystal structure of an iNOS Ca(2+)/CaM-FMN construct, containing the FMN domain in complex with Ca(2+)/CaM, provided structural information on the linkage between the reductase and oxgenase domains of NOS, making it possible to model the holo iNOS structure. This review summarizes recent advances in our understanding of the dynamics of domain movements during CYPOR catalysis and the role of the NOS diflavin reductase domain in the regulation of NOS isozyme activities.
NADPH-细胞色素 P450 氧化还原酶 (CYPOR) 和一氧化氮合酶 (NOS) 是双黄素氧化还原酶家族的两个成员，是含有独特 FAD 和 FMN 结构域的多结构域酶，通过柔性铰链连接。FAD 从 NADPH 接受一个氢原子，还原的 FAD 将电子传递给 FMN，FMN 再将电子传递给细胞色素 P450 或 NOS 加氧酶结构域的血红素中心。该酶家族原型 CYPOR 的结构分析揭示了结构域排列的精确性质以及涉及辅因子结合的残基的作用。最近对 CYPOR 的结构和生物物理研究表明，两个黄素结构域在催化过程中会发生大的结构域运动。NOS 同工酶在还原酶结构域内包含额外的调节元件，通过 Ca(2+)-依赖性钙调蛋白 (CaM) 结合来控制电子传递。最近 iNOS Ca(2+)/CaM-FMN 复合物的晶体结构，包含与 Ca(2+)/CaM 结合的 FMN 结构域，提供了关于 NOS 氧化还原酶结构域和加氧酶结构域之间联系的结构信息，使得构建全酶 iNOS 结构成为可能。本文综述了最近在理解 CYPOR 催化过程中结构域运动的动态以及 NOS 双黄素还原酶结构域在调节 NOS 同工酶活性中的作用方面的进展。
