Identification of the aldolase responsible for the production of 22-hydroxy-23,24-bisnorchol-4-ene-3-one from natural sterols inMycolicibacterium smegmatis

Abstract

The production of 4-androstene-3,17-dione (AD) from natural sterols has been improved by deleting the MSMEG_6561 gene ofMycolicibacterium smegmatiswhich encodes an aldolase. The deletion of this gene increases the AD production yield avoiding the production of 22-hydroxy-23,24-bisnorchol-4-ene-3-one (4-HBC) as by-product and the drawbacks in the AD purification. TheMSMEG_6561gene encodes the only enzyme within theM. smegmatisgenome responsible for the aldolytic activity transforming 22-hydroxy-3-oxo-cholest-4-ene-24-carboxyl-CoA (22-OH-BCN-CoA) into the 4-HBC precursor (20S)-3-oxopregn-4-ene-20-carboxaldehyde (3-OPA). The molar yield of AD production using the MS6039-5941-6561 triple mutant strain was checked in flasks and bioreactor improving very significantly compared with the previously described MS6039-5941 strain.