PROPIONYL-COA ELIMINATION MAY BE A RATE-DETERMINING STEP OF SELECTIVECLEAVAGE OF STEROL SIDE-CHAIN

The effect of propionate on the growth and 4-androstene-3,17-dion (AD) yield was investigated in cultures of Mycobacterium sp. NRRL B-3805 g rowing in minimal medium containing beta-sitosterol as substrate for s elective side chain cleavage. Although the addition of propionate (PA) resulted in a concentration-dependent inhibition of growth at the beg inning of fermentation, cultures started to grow in the presence of 0. 1% of propionic acid reached an AD concentration 38% higher than the c ultures growing in the absence of propionate during two day cultivatio n. After three days of incubation, the AD yields in cultures containin g 0, 0.1 and 0.2% propionate at the inoculation were 68, 79 and 73%, w hile the protein levels were 2.01, 2.11 and 2.60 mg/ml, respectively. Our data showed that the positive effect of PA on the AD production fr om sterols by Mycobacterium sp. NRRL B-3805 could be explained by the induction of the enzymes of the methylmalonate pathway. The activity o f propionyl-CoA carboxylase was about 30% higher in the crude extracts from the induced cultures growing in minimal medium, after 20 hours o f growth, than in those from the controls (18.2 and 14.1 mU/mg, respec tively, using propionyl-CoA as substrate). The distribution of the aci d-stable C-14-radioactivity which built into methylmalonate, succinate and fumarate indicated that methylmalonyl-CoA mutase was also induced . Our data demonstrated that elimination of the toxic propionyl-CoA re leased from the side chain of the sterol is likely the rate-determinin g step of the AD production, at least at the beginning of the process.