A Keto Reductase Involved in Steroid Degradation in Mycolicibacterium neoaurum

Phytosterols can be used by microorganisms as carbon and energy sources and completely degraded into CO₂ and H₂ O. The catabolic pathway of phytosterols was well characterized in many microorganisms. Blocking the steroid core ring degradation by deletions of fadE30 and fadD3 genes, two important steroid intermediates, 3aα-H-4α-(3'-Propionic acid)-5α-hydroxy-7aβ-methylhexahydro-1-indanone-δ-lactone (sitolactone, or HIL) and 3aα-H-4α-(3'-propionic acid)-7aβ-methylhexahydro-1,5-indanedione (HIP) can be accumulated. They are currently used to synthesize nor-steroid drugs with an α-methyl group or without the methyl group at the C₁₀ -position, such as estrone and norethindrone. In this study, a key gene involved in the bioconversion of HIP to HIL was identified in Mycolicibacterium neoaurum. Through heterologous expression, gene hipR was found to be involved in the reduction of the C₅ keto group of HIP to a hydroxy group, leading to spontaneously lactonization into HIL in vitro. Through gene complementation and knockout, HipR functions were verified and two HIP degradation pathways in vivo were elucidated. The finding of this research facilitated the understanding of the metabolic pathway of sterols, and was directly applied to engineering robust production strains by overexpression or knockout of related genes.