Stereoselective carveol dehydrogenase from Rhodococcus erythropolis DCL14 -: A novel nicotinoprotein belonging to the short chain dehydrogenase/reductase superfamily
Stereoselective carveol dehydrogenase from Rhodococcus erythropolis DCL14 -: A novel nicotinoprotein belonging to the short chain dehydrogenase/reductase superfamily
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DOI：
10.1074/jbc.274.37.26296
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发表时间：
1999-09-10
影响因子：
4.8
通讯作者：
van Berkel, WJH
中科院分区：
文献类型：
--
作者：
van der Werf, MJ;van der Ven, C;van Berkel, WJH
一种新型的烟毒素，催化摄影酚二氯苯丁醇依赖性的氧化为葡萄干，被纯化为赤型甲状腺癌DCL14的均匀性。该酶在柠檬烯和甲状腺二氯苯酚依赖性的脱氢酶（CDH）上的生长后特异性诱导是120 kDa的同倍化合物，每个亚基含有紧密结合的NAD（H）分子。该酶在pH 5.5和50摄氏度下具有最佳活性，并显示出较大的底物特异性，偏爱取代的环己醇。当与（4R） - 或（4S） - 碳水化合物的非对映异构体混合物孵育时，CDH立体选择性地催化了（6s） - 碳水化合物立体异构体的转化，具有纯立体异构体的动力学研究表明，这是由于V（最大差异） ）/k（m）值和同时抑制cr） - 或（S） - 瓦隆。在编码涉及二烯烯降解的酶的操纵子中鉴定了红菌CDH基因（LIMC）。 CDH核苷酸序列揭示了831个碱基对的开放式阅读框，编码了277-氨基酸蛋白，推导质量为29,531 Da。 CDH初级结构与短链脱氢酶/还原酶超家族的成员共享10-30％的序列认同。结构同源性与三二硫代乙烯还原酶fi om magnaporthe grisea提示 - 来自r。erythropolisdcl14的CDH是α/β的一域蛋白质，具有额外的回路插入，涉及NAD结合，并且涉及柔性的C-终端c-terminal涉及单纤维结合。
A novel nicotinoprotein, catalyzing the dichlorophenolindophenol-dependent oxidation of carveol to carvone, was purified to homogeneity from Rhodococcus erythropolis DCL14. The enzyme is specifically induced after growth on limonene and carveol Dichlorophenolindophenol-dependent carveol dehydrogenase (CDH) is a homotetramer of 120 kDa with each subunit containing a tightly bound NAD(H) molecule. The enzyme is optimally active at pH 5.5 and 50 degrees C and displays a broad substrate specificity with a preference for substituted cyclohexanols. When incubated with a diastereomeric mixture of (4R)- or (4S)-carveol, CDH stereoselectively catalyzes the conversion of the (6S)-carveol stereoisomers only, Kinetic studies with pure stereoisomers showed that this is due to large differences in V(max)/K(m) values and simultaneous product inhibition by CR)- or (S)-carvone. The R. erythropolis CDH gene (limC) was identified in an operon encoding the enzymes involved in Iimonene degradation. The CDH nucleotide sequence revealed an open reading frame of 831 base pairs encoding a 277-amino acid protein with a deduced mass of 29,531 Da. The CDH primary structure shares 10-30% sequence identity with members of the short chain dehydrogenase/reductase superfamily. Structure homology modeling with trihydroxynaphthalene reductase fi om Magnaporthe grisea suggests-that CDH from R. erythropolis DCL14 is an alpha/beta one-domain protein with an extra loop insertion involved in NAD binding and a flexible C-terminal part involved in monoterpene binding.