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- Title
Engineering a hydroxysteroid dehydrogenase to improve its soluble expression for the asymmetric reduction of cortisone to 11β-hydrocortisone.
- Authors
Zhang, Dalong; Zhang, Rui; Zhang, Jie; Chen, Liying; Zhao, Chunxia; Dong, Wenyue; Zhao, Qing; Wu, Qiaqing; Zhu, Dunming
- 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.
- Subjects
HYDROXYSTEROID dehydrogenases; ALCOHOL dehydrogenase; CORTISONE; BIOCATALYSIS; HYDROCORTISONE regulation; MUTAGENICITY testing
- Publication
Applied Microbiology & Biotechnology, 2014, Vol 98, Issue 21, p8879
- ISSN
0175-7598
- Publication type
Article
- DOI
10.1007/s00253-014-5967-1