Cu(II)‐enhanced microbial Cr(VI) reduction is common in the environment, yet its mechanism is unknown. The specific activity of chromate reductase, NfoR, from Staphylococcus aureus sp. LZ‐01 was augmented 1.5‐fold by Cu(II). Isothermal titration calorimetry and spectral data show that Cu(II) binds to NfoR nonspecifically. Further, Cu(II) stimulates the nitrobenzene reduction of NfoR, indicating that Cu(II) promotes electron transfer. The crystal structure of NfoR in complex with CuSO₄ (1.46 Å) was determined. The overall structure of NfoR‐Cu(II) complex is a dimer that covalently binds with FMN and Cu(II)‐binding pocket is located at the interface of the NfoR dimer. Structural superposition revealed that NfoR resembles the structure of class II chromate reductase. Site‐directed mutagenesis revealed that Leu⁴⁶ and Phe¹²³ were involved in NADH binding, whereas Trp⁷⁰ and Ser⁴⁵ were the key residues for nitrobenzene binding. Furthermore, His¹⁰⁰ and Asp¹⁷¹ were preferential affinity sites for Cu(II) and that Cys¹⁶³ is an active site for FMN binding. Attenuation reductase activity in C163S can be partially restored to 54% wild type by increasing Cu(II) concentration. Partial restoration indicates dual‐channel electron transfer of NfoR via Cu(II) and FMN. We propose a catalytic mechanism for Cu(II)‐enhanced NfoR activity in which Cu(I) is formed transiently. Together, the current results provide an insight on Cu (II)‐induced enhancement and benefit of Cr(VI) bioremediation.

中文翻译：

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铜（II）非特异性结合铬还原酶NfoR促进Cr（VI）还原

Cu（II）增强微生物中的Cr（VI）还原在环境中很常见，但其机理尚不清楚。来自金黄色葡萄球菌的铬酸还原酶NfoR的比活性。LZ -01被Cu（II）增强了1.5倍。等温滴定热法和光谱数据表明，Cu（II）与NfoR非特异性结合。此外，Cu（II）刺激NfoR的硝基苯还原，表明Cu（II）促进电子转移。NfoR与CuSO ₄配合物的晶体结构（1.46Å）被确定。NfoR-Cu（II）复合物的整体结构是与FMN共价结合的二聚体，而Cu（II）结合口袋位于NfoR二聚体的界面。结构重叠表明NfoR类似于II类铬酸还原酶的结构。定点诱变表明Leu ⁴⁶和Phe ¹²³参与了NADH的结合，而Trp ⁷⁰和Ser ⁴⁵是与硝基苯结合的关键残基。此外，His ¹⁰⁰和Asp ¹⁷¹是Cu（II）和Cys ^163的优先亲和位点是FMN绑定的活动站点。通过增加Cu（II）浓度，C163S中的衰减还原酶活性可以部分恢复为54％的野生型。部分恢复表明NfoR通过Cu（II）和FMN的双通道电子转移。我们提出了Cu（II）增强NfoR活性的催化机制，其中Cu（I）瞬时形成。在一起，当前的结果提供了对Cu（II）诱导的Cr（VI）生物修复的增强和益处的见解。