title: Escherichia coli kduD encodes an oxidoreductase that converts both sugar and steroid substrates.
abstract: A previously unidentified oxidoreductase from Escherichia coli catalyzes the regioselective reduction of eukaryotic steroid hormone 11-deoxycorticosterone (11-DOC) to the valuable bioactive product 4-pregnen-20,21-diol-3-one. In nature, a reduction of C-20 carbonyl of C21 steroids is catalyzed by diverse NAD(P)H-dependent oxidoreductases. Enzymes that possess 20-ketosteroid reductase activity, however, have never before been described in E. coli. Our present study aimed to identify and characterize the E. coli enzyme which possesses 20-ketosteroid reductase activity against eukaryotic steroid hormone 11-DOC. We partially purified the enzyme from E. coli DH5α using protein chromatography techniques. Mass spectrometry revealed the presence of three NADH-specific oxidoreductases in the sample. The genes encoding these oxidoreductases were cloned and overexpressed in E. coli UT5600 (DE3). Only the overexpression of 2-dehydro-3-deoxy-D-gluconate 5-dehydrogenase (KduD) encoded by kduD gene enabled the whole-cell biotransformation of 11-DOC. A 6xHis-tagged version of KduD was purified to homogeneity and found to reduce several eukaryotic steroid hormones and catalyze the conversion of novel sugar substrates. KduD from E. coli is therefore a promiscuous enzyme that has a predicted role in sugar conversion in vivo but can be used for the production of valuable bioactive 20-hydroxysteroids.
authors: Institut für Pharmazeutische und Medizinische Chemie, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany.
authors: Agne
authors: A
authors: Tubeleviciute
authors: Mark George
authors: MG
authors: Teese
authors: Joachim
authors: J
authors: Jose
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001711:Biotransformation; D050260:Carbohydrate Metabolism; D003001:Cloning, Molecular; D003900:Desoxycorticosterone / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D015394:Molecular Structure; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D003900:Desoxycorticosterone; D010088:Oxidoreductases; C093584:gluconate 5-dehydrogenase