Microbial transformation of aklanonic acid, a potential early intermediate in the biosynthesis of anthracyclines.
Sir: Recently, we isolated a new pigment from fermentations of Streptomyces sp. ZIMET 43717". The close structural relationship of this compound (aklanonic acid, Ia, Fig. 1) to anthracyclinones induced us to attempt its biotransformation to anthracyclines by daunorubicin-negative mutants derived from different daunorubicin producing organisms. The results of our experiments provided evidence that aklanonic acid very probably is an early intermediate in the biosynthetic pathway of anthracyclines. Thus, aklanonic acid, its methyl ester, and aklanone, a chemical conversion product of aklanonic acid (Fig. 1)", were fed to fermentations of the mutant strains O3P; and IP,. Both mutant strains were isolated after N-methyl-N'nitro-N'-nitrosoguanidine treatment of daunorubicin-producing Streptomyces griseaus strains. Strain O1P; was derived from IMET JA 5142. Strain IP, is a negative mutant of strain 1P which produces a mixture of new red and blue pigments"'. The chemical structures of the two main red components designated 1P/I and IP/II3) are shown in Fig. 1. Fermentations of the strains O1P; and 1P, were carried out as described2) in 500-m1 flasks each containing 80 ml of medium for 24 hours. Then 3 ml of a solution of substrate in methanol (1 mg/ml) were added to each flask and cultivation continued for 24 hours. Extracts of mycelium with acetone and chloroform (1: 1) were purified by chromatography on KH2PO4 buffered silica gel (chloroform acetone, 10: 2). Identification of the biotransformation products was performed as indicated in Table 1. Glycosides extracted from the culture filtrate were hydrolyzed with 0.1 N HCI. Compound IP/II was purified by column chromatography on silica gel with benzene. The results are listed in Table 1. Aklanonic acid as well as its methyl ester were transformed by mutant O,P, to give e-rhodomycinone, 7deoxy-e-rhodomycinone and glycosides of dauno-
BIOSYNTHESIS OF DAUNORUBICIN GLYCOSIDES: ANALYSIS WITH BLOCKED MUTANTS
ABSTRACT Daunorubicin (daunomycin; NSC 82151) is an anthracycline antibiotic that is useful in the treatment of human leukemias. Several Streptomyces species accumulate higher glycosides of daunorubicin, which can be converted to daunorubicin by simple hydrolysis. We isolated a number of mutants of a daunorubicin glycoside-producing streptomycete, and found that some were deficient in accumulation of daunorubicin glycosides. One such mutant accumulated no detectable anthracyclines, while other mutants accumulated anthracyclines or aglycones that have not been reported before or have been described as co-metabolites of daunorubicin glycosides. Some of these anthracyclines and aglycones were purified and used as substrates in bioconversion experiments with various mutants. The results allowed us to construct a proposed pathway for the biosynthesis of daunorubicin glycosides by this streptomycete. KEYWORDS Anthracycline; daunorubicin; biosynthesis; blocked mutants; biotransformation.
Detection and study of cosynthesis of tetracycline antibiotics by an agar method.
SUMMARY: Pairs of non-tetracycline-producing mutants of Streptomyces rimosus or S. aureofaciens were grown side by side on agar. Their ability to produce antibiotic by cosynthesis was tested by placing a strip of agar cut from the combined culture on plates containing Bacillus subtilis. The activity was revealed as an inhibition halo formed on B. subtilis, opposite one or other mutant strain. The strain surrounded by the halo was considered as a converter of an intermediate product secreted by the other strain. Two types of mutants were observed: a rare type probably affecting the main pathway of antibiotic biosynthesis, and a more frequent type probably affecting some regulatory process.