Actinomycete cytochrome P450 from Pseudonocardia autotrophica that catalyzes vitamin D3 hydroxylation

摘要

Pseudonocardia autotropica was known as an industrially important actinomycete strain that is capable of hydroxylating vitamin D3 (VD3) to 1alpha,25-dihydroxy VD3 (calcitriol) in a similar way of human CYP27-mediated metabolic system. For more advantageous bioprocess, the responsible hydroxylase (Vdh) was purified and characterized. The Vdh was identified to be the novel cytochrome P450 and catalyzed 25- and sequential 1alpha-hydroxylations for VD3 in reconstitution system. The gene for Vdh was expressed in Escherichia coli and Rhodococcus erythropolis, and provided whole-cell biocatalysts in hydroxylation of VD3. Directed evolution enabled us to obtain the improved Vdh that showed a 21-fold increased activity in 25-hydroxlation for VD3, indicating a promising approach to the refined bioprocess for the hydroxyl VD3.