Enhancing the production of physiologically active vitamin D3 by engineering the hydroxylase CYP105A1 and the electron transport chain

In this study, the conversion of vitamin D-3 (VD3) to its two active forms 25(OH)VD3 and 1 alpha, 25(OH)(2)VD3 was carried out by engineering the hydroxylase CYP105A1 and its redox partners Fdx and Fdr. CYP105A1 and Fdx-Fdr were respectively expressed in E. coli BL21(DE3) and purified. The electron transport chain Fdx-Fdr had higher selectivity for the coenzyme NADH than NADPH. HPLC analysis showed that CYP105A1 could hydroxylate the C25 and C1 alpha sites of VD3 and convert VD3 to its active forms. Finally, a one-bacterium-multi-enzyme system was constructed and used in whole-cell catalytic experiments. The results indicated that 2.491 mg/L of 25(OH)VD3 and 0.698 mg/L of 1 alpha, 25(OH)(2)VD3 were successfully produced under the condition of 1.0% co-solvent DMSO, 1 mM coenzyme NADH and 35 g/L biocatalyst loading. This study contributes to a basis for the industrial production of active VD3 in future.

Automated summary

In this study, vitamin D-3 (VD3) was converted to its active forms 25(OH)VD3 and 1 alpha, 25(OH)(2)VD3 by engineering CYP105A1 and its redox partners. A one-bacterium-multi-enzyme system was constructed and successfully produced active VD3.