State Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Collaborative Innovation Center for Green Transformation of Bio-Resources, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan, People's Republic of China.
Department of Thoracic Surgery, Tongji Hospital, Tongji Medical Collage of Huazhong University of Science and Technology, Wuhan, People's Republic of China.
Appl Environ Microbiol. 2023 Mar 29;89(3):e0196322. doi: 10.1128/aem.01963-22. Epub 2023 Feb 28.
In this study, a new cytochrome P450 enzyme, namely, CYP68J5_Fusarium graminearum (CYP68J5_fg), was identified from Fusarium graminearum via a combination of transcriptome sequencing and heterologous expression in Saccharomyces cerevisiae. The biotransformation of progesterone by whole-cells of S. cerevisiae expressing CYP68J5_fg revealed that the CYP68J5_fg possessed steroidal 12β- and 15α-hydroxylase activities. To the best of our knowledge, this is the first time that a fungal P450 enzyme with 12β-hydroxylase activity has been identified. This advance offers opportunities to boost the efficiency and selectivity of the CYP68J5_fg hydroxylating system and thus shows great potential for further applications of this enzyme for the synthesis of steroid drugs. Regioselective and stereoselective hydroxylation is of vital importance in the functionalization of steroids, which remains challenging in organic synthesis. In particular, the C12-hydroxy steroids play a significant role in the synthesis of many important steroidal drugs. In this study, a novel fungal P450 enzyme with 12β-hydroxylation activity was identified, and it shows different substrate specificity and regioselectivity, compared to the bacterial and fungal steroidal hydroxylases that are known to date. This lays the foundation for the creation of effective biocatalysts for the process of 12β-hydroxylation, although further understanding of the molecular structural basis of this fungal P450 is needed to facilitate the engineering of this enzyme for industrial applications.
在这项研究中，通过转录组测序和在酿酒酵母中的异源表达相结合，从禾谷镰刀菌中鉴定出一种新的细胞色素 P450 酶，即 CYP68J5_Fusarium graminearum（CYP68J5_fg）。通过表达 CYP68J5_fg 的酿酒酵母全细胞对孕酮的生物转化表明，CYP68J5_fg 具有甾体 12β-和 15α-羟化酶活性。据我们所知，这是首次鉴定出具有 12β-羟化酶活性的真菌 P450 酶。这一进展为提高 CYP68J5_fg 羟化系统的效率和选择性提供了机会，从而为该酶在甾体药物合成中的进一步应用展示了巨大的潜力。在甾体的功能化中，区域和立体选择性羟化至关重要，这在有机合成中仍然具有挑战性。特别是 C12-羟基甾体在许多重要甾体药物的合成中起着重要作用。在这项研究中，鉴定出了一种具有 12β-羟化活性的新型真菌 P450 酶，与已知的细菌和真菌甾体羟化酶相比，它表现出不同的底物特异性和区域选择性。这为创建有效的 12β-羟化生物催化剂奠定了基础，尽管需要进一步了解这种真菌 P450 的分子结构基础，以促进该酶在工业应用中的工程改造。
