Functional characterization of 3-Ketosteroid 9α-Hydroxylases in Rhodococcus ruber strain Chol-4.

•Three KshAs and just one KshB subunit have been identified in Rhodococcus ruber.•KshA2 isoform is involved in the degradation of steroid substrates with a short side chain in their molecule.•KshA3 is involved in the degradation of steroids with long side chains.•KshA1 is a more versatile enzyme that is related to the cholic acid catabolism.•The steroid side chain degradation is independent of KshAB activities in this strain.