Bacterial metabolism of anthracycline antibiotics. Steffimycinone and steffimycinol conversions.

Streptomyces nogalater, UC-2783, and Streptomyces peucetius var. caesius, IMRU-3920/UC-5633, catalyze ketonic carbonyl reduction of steffimycinone (1, Scheme 1). Using cell-free preparations of S. nogalater, the process of ketonic carbonyl reduction has been shown to be TPNH linked. The product, steffimycinol (2), is reduced further by Aeromonas hydrophila, 2C/UC-6303, by the process of microaerophilic conversion of anthracyclinones previously reported1,2) with the result being the formation of 7-deoxysteffimycinol (3). The products (2 and 3) were isolated by extraction from the fermentations followed by chromatographic purification. Identification was by comparison of various physical properties and spectral data with those of authentic materials obtained by chemical means. Catalytic activity of the crude enzyme preparations of S. nogalater was lost by dialysis by restored by addition of TPNH although not by addition of DPNH demonstrating TPNH dependence. The reaction rate increased linearly with added crude enzyme protein up to 4 mg/ml and was highest between pH 6.5 and 7.0.

Reductive microbial conversion of anthracycline antibiotics.

Reductive conversion of several anthracycline glycosides to their 7-deoxyaglycones occurs during their microaerophilic incubation with strains of Aeromonas hydrophila, Citrobacter freundii, and Escherichia coli. Further, extracts of microaerophilically grown A. hydrophilia catalyze DPNH-dependent reductive conversion of the same compounds. Anthracycline substrates cleaved by both whole cells and by the cell-free system include steffimycin, steffimycin B, nogalamycin, cinerubin A, and daunomycin. Investigation of glycoside cleavage as a function of both time and anthracycline concentration demonstrated the superiority of A. hydrophila over C. freundii and E. coli in regard to reaction rate and efficiency of conversion. Interestingly, some degree of anaerobicity was required for glycoside cleavage by all three organisms. Evidence supporting 7-deoxyaglycone formation via direct reductive cleavage, as opposed to a multienzyme-catalyzed process involving hydrolysis followed by dehydration and reduction, includes the following. Equilibrium mixtures of glycoside substrate and 7-deoxyaglycone product prepared using both whole cells and their extracts display no anthracycline hydrolysis products. Further, authentic steffimycinone (aglycone), the expected product of hydrolytic sugar cleavage of steffimycin, was shown to be converted to 7-deoxysteffimycinone (7-deoxyaglycone) at a rate slower than steffimycin. These data indicate that, if steffimycinone were present as an unbound metabolic intermediate, it should have been visible in the equilibrium mixture, but none was detected.