Synthetic glucocorticoids such as methylprednisolone are compounds of fundamental interest to the pharmaceutical industry as their modifications within the sterane scaffold lead to higher inflammatory potency and reduced side effects compared with their parent compound cortisol. In methylprednisolone production, the complex chemical hydroxylation of its precursor medrane in position C21 exhibits poor stereo‐ and regioselectivity making the process unprofitable and unsustainable. By contrast, the use of a recombinant E. coli system has recently shown high suitability and efficiency. In this study, we aim to overcome limitations in this biotechnological medrane conversion yielding the essential methylprednisolone‐precursor premedrol by optimizing the CYP21A2‐based whole‐cell system on a laboratory scale. We successfully improved the whole‐cell process in terms of premedrol production by (a) improving the electron supply to CYP21A2; here we use the N‐terminally truncated version of the bovine NADPH‐dependent cytochrome P450 reductase (bCPR₋₂₇) and coexpression of microsomal cytochrome b₅; (b) enhancing substrate access to the heme by modification of the CYP21A2 substrate access channel; and (c) circumventing substrate inhibition which is presumed to be the main limiting factor of the presented system by developing an improved fed‐batch protocol. By overcoming the presented limitations in whole‐cell biotransformation, we were able to achieve a more than 100% improvement over the next best system under equal conditions resulting in 691 mg·L⁻¹·d⁻¹ premedrol.

中文翻译：

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使用重组大肠杆菌鉴定和规避CYP21A2介导的前药生产中的瓶颈。

合成的糖皮质激素（例如甲基强的松龙）是制药行业最感兴趣的化合物，因为与它们的母体化合物皮质醇相比，它们在甾烷骨架中的修饰导致更高的炎症效力和更低的副作用。在甲基强的松龙生产中，其前体甲乙炔在位置C21的复杂化学羟基化显示出较差的立体选择性和区域选择性，使该过程无利可图且不可持续。相比之下，重组大肠杆菌的使用该系统最近显示出很高的适用性和效率。在这项研究中，我们旨在通过在实验室规模上优化基于CYP21A2的全细胞系统来克服这种生物技术甲基溴转化产生必需的甲基强的松龙前体前药的局限性。通过（a）改善CYP21A2的电子供应，我们成功地改善了全细胞过程的前药生产。在这里，我们使用牛NADPH依赖性细胞色素P450还原酶（bCPR ₋₂₇）的N末端截短版本和微粒体细胞色素b _5的共表达; （b）通过修饰CYP21A2底物进入通道，增加底物进入血红素的途径；（c）通过开发改进的补料分批操作规程来规避底物抑制，这被认为是本系统的主要限制因素。通过克服目前在全细胞生物转化中的局限性，我们在同等条件下比次优系统获得了100％的改善，可产生691 mg·L ^-1 ·d ^-1的前药。