1996 Volume 10 Issue 1 Pages 1-11
Streptomyces cholesterol oxidase, an enzyme on the first step of microbial catabolism of cholesterol, was overproduced in a secretory fashion in Streptomyces lividans with a multi-copy shuttle vector. The gene coding for cholesterol oxidase (choA) was modified and overexpressed in Escherichia coli. The modified choA gene was also used to develop promoter-probe vectors to use in enteric bacteria. Expression of the choA gene in lactic acid bacteria might facilitate the degradation of cholesterol in dairy food. The Streptomyces cholesterol oxidase was unexpectedly found to constitute a novel class of insecticidal proteins that may be useful for controlling pests that are resistant to Bacillus thuringiensis toxins.
Furthermore, the Arthrobacter simplex gene (ksdD) encoding 3-ketosteroid-Δ1-dehydrogenase, a key enzyme in the degradation of the steroid nucleus, was cloned and overexpressed in S. lividans using a multi-copy shuttle vector, leading to an about 100-fold overproduction of the enzyme compared to the natural producer. Nucleotide sequence analysis revealed that ksdD is clustered with two more genes possibly involved in steroid catabolism. Upstream of ksdD, a gene (ksdR) encoding a hypothetical regulatory protein that shows similarities to several regulators was found. Adjoining downstream to ksdD in an apparent translational coupling is a gene (ksdI) coding for a protein that would display strong similarities to 3-ketosteroid-Δ5-isomerase. Further sequence analysis downstream of ksdI revealed three ORFs. The deduced protein product of one of these showed significant similarities to the KsdD protein. Thus, genes involved in steroid metabolism seem to be clustered in bacteria.
