Abstract
This study aims to improve the efficiency of 4-androstene-317-dione (AD) production from high-concentrated phytosterols through a combined strategy of metabolic pathway regulation and two-step bioprocess. Firstly, we overexpressed cholesterol oxidase and steroid C27 monooxygenase to strengthen the metabolic flux to AD and then enhanced the expression of 17β-hydroxysteroid dehydrogenase to reduce the by-product 20-hydroxy-methylpregna-4-ene-3-one. As a result, the AD production was enhanced from 2.4 to 4.5 g/L with an increase of 88.6 %. Afterward, a two-step bioprocess, cell culture at 30 °C and bioconversion with resting cells at 37 °C, was adopted to avoid the cell growth inhibition and decrease the steroid nucleus degradation. Finally, 25.8 g/L AD was produced from 50 g/L phytosterols, which is significantly higher than that from the fermentation bioprocess and is the highest reported AD production. Therefore, this work hopes to pave the way for the commercial production of AD through Mycobacterium transformation.
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