delCardayre S B, Davies J E
Department of Microbiology and Immunology, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
J Biol Chem. 1998 Mar 6;273(10):5752-7. doi: 10.1074/jbc.273.10.5752.
The cdr gene encoding coenzyme A disulfide reductase (CoADR) from Staphylococcus aureus 8325-4 was cloned, sequenced, and overexpressed. The gene encodes a 438-amino acid polypeptide that has a calculated molecular weight of 49,200 and sequence similarity to the pyridine nucleotide-disulfide oxidoreductase family of flavoenzymes. The deduced primary structure contains consensus sequences for flavin adenine dinucleotide and NADPH-binding regions but lacks the catalytic disulfide signature sequence typical of the glutathione reductase family of disulfide reductases. The active site region of CoADR has only a single cysteine residue that is similar to that in the conserved SFXXC active site motif of NADH oxidase and NADH peroxidase from Enterococcus faecalis. CoADR is the first disulfide reductase reported having this active site region, and sequence comparisons of CoADR to representative members of the pyridine nucleotide-disulfide reductase superfamily placed CoADR in a distinct subfamily. CoADR was overexpressed in Escherichia coli using the pET expression system, and 5-10 mg of fully active recombinant enzyme were recovered per liter of E. coli cells.
克隆、测序并过量表达了来自金黄色葡萄球菌8325 - 4的编码辅酶A二硫化物还原酶（CoADR）的cdr基因。该基因编码一个438个氨基酸的多肽，其计算分子量为49,200，并且与黄素酶的吡啶核苷酸 - 二硫化物氧化还原酶家族具有序列相似性。推导的一级结构包含黄素腺嘌呤二核苷酸和NADPH结合区域的共有序列，但缺乏二硫化物还原酶谷胱甘肽还原酶家族典型的催化二硫键特征序列。CoADR的活性位点区域只有一个半胱氨酸残基，类似于粪肠球菌NADH氧化酶和NADH过氧化物酶保守的SFXXC活性位点基序中的半胱氨酸残基。CoADR是报道的第一个具有该活性位点区域的二硫化物还原酶，并且CoADR与吡啶核苷酸 - 二硫化物还原酶超家族的代表性成员的序列比较将CoADR置于一个独特的亚家族中。使用pET表达系统在大肠杆菌中过量表达CoADR，每升大肠杆菌细胞可回收5 - 10毫克完全活性的重组酶。
