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Updated: Sep 29, 2025
A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products
Published on: October 4, 2019
A New 3-Ketosteroid-Δ1-Dehydrogenase with High Activity and Broad Substrate Scope for Efficient Transformation of
Yu Wang1, Rui Zhang1, Jinhui Feng1
1National Technology Innovation Center of Synthetic Biology, National Engineering Laboratory for Industrial Enzymes, Tianjin Engineering Research Center of Biocatalytic Technology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin Airport Economic Area, 32 Xi Qi Dao, Tianjin 300308, China.
Abstract:
3-Ketosteroid-Δ1-dehydrogenases (KstDs [EC 1.3.99.4]) catalyze the Δ1-dehydrogenation of steroids and are a class of important enzymes for steroid biotransformations. In this study, nine putative kstD genes from different origins were selected and overexpressed in Escherichia coli BL21(DE3). These recombinant enzymes catalyzed the Δ1-desaturation of a variety of steroidal compounds. Among them, the KstD from Propionibacterium sp. (PrKstD) displayed the highest specific activity and broad substrate spectrum. The detailed catalytic characterization of PrKstD showed that it can convert a wide range of 3-ketosteroid compounds with diverse substituents, ranging from substituents at the C9, C10, C11 and C17 position through substrates without C4-C5 double bond, to previously inactive C6-substituted ones such as 11β,17-dihydroxy-6α-methyl-pregn-4-ene-3,20-dione. Reaction conditions were optimized for the biotransformation of hydrocortisone in terms of pH, temperature, co-solvent and electron acceptor. By using 50 g/L wet resting E. coli cells harboring PrKstD enzyme, the conversion of hydrocortisone was about 92.5% within 6 h at the substrate concentration of 80 g/L, much higher than the previously reported results, demonstrating the application potential of this new KstD.
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