title: Engineering a hydroxysteroid dehydrogenase to improve its soluble expression for the asymmetric reduction of cortisone to 11β-hydrocortisone.
abstract: 11β-Hydrocortisone (11β-HC) is an important anti-inflammatory drug and intermediate for the synthesis of other steroids. One of the methods for the synthesis of 11β-HC is the asymmetric reduction of cortisone catalyzed by a highly regioselective and stereoselective 11β-hydroxysteroid dehydrogenase (11β-HSDH). However, this process has been prohibited by the poor soluble expression of the membrane-anchoring protein 11β-HSDH in prokaryotes. To overcome this obstacle, a mutant III-1G1 (Phe80Leu/Thr105Ser/Ala260Thr/Tyr274Stop) truncated at position 274 with improved yield of soluble protein was stepwise obtained from the 11β-HSDH from guinea pig by random mutagenesis combining with structural complementation assay and C-terminal truncating library screening. The improved 11β-HSDH mutant and glucose dehydrogenase (GDH) from Bacillus subtilis were coexpressed in Escherichia coli. The resulting whole-cell biocatalyst catalyzed the reduction of cortisone to 11β-HC with 98 % conversion in 20 h, laying foundation for the development of an asymmetric reduction process for the production of 11β-HC.
authors: National Engineering Laboratory for Industrial Enzymes and Tianjin Engineering Center for Biocatalytic Technology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China.
authors: Dalong
authors: D
authors: Zhang
authors: Rui
authors: R
authors: Zhang
authors: Jie
authors: J
authors: Zhang
authors: Liying
authors: L
authors: Chen
authors: Chunxia
authors: C
authors: Zhao
authors: Wenyue
authors: W
authors: Dong
authors: Qing
authors: Q
authors: Zhao
authors: Qiaqing
authors: Q
authors: Wu
authors: Dunming
authors: D
authors: Zhu
mesh_terms: D000818:Animals; D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics; D001711:Biotransformation; D003001:Cloning, Molecular; D003348:Cortisone / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D042843:Glucose 1-Dehydrogenase / Q000235:genetics / Q000378:metabolism; D006168:Guinea Pigs / Q000235:genetics; D006854:Hydrocortisone / Q000378:metabolism*; D006913:Hydroxysteroid Dehydrogenases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010084:Oxidation-Reduction; D015202:Protein Engineering; D017384:Sequence Deletion; D012995:Solubility
chemical_list: D050505:Mutant Proteins; D006913:Hydroxysteroid Dehydrogenases; D042843:Glucose 1-Dehydrogenase; D003348:Cortisone; D006854:Hydrocortisone