Steroid-1-dehydrogenase of Rhodococcus erythropolis: purification and N-terminal amino acid sequence.

The inducible steroid-1-dehydrogenase from the bacterium Rhodococcus erythropolis IMET 7030 was purified to homogeneity using affinity chromatographic, electrophoretic, and ion exchange techniques. The spectrum of the pure enzyme is characterized by the associated FAD. The M(r) of the enzyme is 56,000.

Localization of the cholesterol oxidase in Rhodococcus erythropolis IMET 7185 studied by immunoelectron microscopy.

Rhodococcus erythropolis IMET 7185 produces an inducible cholesterol oxidase (COD) which can easily be extracted by treatment of cells with 0.1% Triton X-100. The yield of the enzyme was 3.

Localization of the steroid 1-dehydrogenase in Rhodococcus erythropolis IMET 7030 by immunoelectron microscopy.

The steroid 1-dehydrogenase of Rhodococcus erythropolis IMET 7030, an active steroid-transforming strain, was localized by immunogold labelling both in cells induced with 17-alpha-methyl-testosterone and in noninduced cells. The labelling intensity was much higher in induced cells than in noninduced cells, indicating increased enzyme production in the case of induction. Using the postembedding procedure, the main portion of the enzyme was found in the peripheral region of the cytoplasm.

Application of newly synthesized detergents in the side chain degradation of plant sterols by Mycobacterium fortuitum.

Newly synthesized detergents of polyoxyethylene polyoxypropylene co-polymers (Co-EOPO), polyoxyethylene polyoxypropylene adducts of the diethylene triamine (DETA-EOPO), and steroidal detergents (SDD) stimulate significantly side chain degradation of plant sterols referring to both the sterol degradation and the formation of the product, 9 alpha-hydroxy-androsta-4-ene-3,17-dione (9 OH-AD), by Mycobacterium fortuitum NRRL-B-8119. Highest stimulative effects were observed with derivatives of the DETA-EOPO group. Using compounds of the Co-EOPO group optimal sterol transformation rates were found for polymers with a polyoxypropylene domain of 1400-2000 in molecular weight and with a polyoxyethylene content of 20-50%.

Overexpression of a Rhodococcus erythropolis protein in Escherichia coli with immunological identity to the Rhodococcus steroid 1-dehydrogenase. Immunoelectron microscopic localization and electrophoretic studies.

The recombinant Escherichia coli K-12 strain chi 6060 harbouring the plasmid pYA 1201 with a gene from Rhodococcus erythropolis IMET 7030 overexpressed a protein which reacts with a monospecific antiserum against the steroid 1-dehydrogenase (Sdh) from the same Rhodococcus strain. It was shown previously that this recombinant protein exhibits no enzymatic activity. By immunogold labelling the protein was localized on ultrathin sections of the recombinant E.