It is the objective of the proposed research to advance kowledge on the biosynthesis of the antitumor antibiotic mitomycin, a product of the fermentations of several Streptomyces species including S. michiganesis. It is also the objective of the proposed research to investigate the biosynthetic origin of michigazone, a pigment which is a co-metabolite of mitomycin in S. michiganesis. The pigment and mitomycin appear to share the common presence of a meta-C-C6-N unit in a portion of their structures which however, shows differences in the two compounds. Related meta-C-C6-N units occur in the ansamycin antibiotics, including rifamycin and antitumor antibiotic ansamitocin, as well as in other antibiotics. All of these meta-C-C6-N units appear to be biosynthetically derived from a novel branch of the shikimic acid pathway. In addition to the studies on the origin of the meta-C-C6-N units, the role D-glucosamine plays in mitomycin biosynthesis will be further investigated and the biosynthetic origin of the second ring of michigazone will be examined. The goals will be pursued in feeding experiments with radioactively or stable isotope labeled precursors, several of which will be synthesized, and by radioactivity and 13C NMR analysis. It is also planned to isolate mitomycin and michigazone nonproducing mutants as well as auxotrophic mutants from the strains of interest and to develop genetic studies in these organisms. Particular emphasis will be placed in the genetic studies on attempts to establish in our strains cloning vectors developed in D. A. Hopwood's laboratory and elsewhere. Once this is accomplished it will be explored if mitomycin and michigazone nonproducing mutants can be complemented by cloned DNA. This would constitute the most direct access to genes involved in the elaboration of these compounds. Once cloned pathway genes are available they can be used as probes to determine the distribution of the genes required for meta-C-C6-N unit biosynthesis among members of the genus Streptomyces and to examine the regulation of their expression. Genetic information on the determination of the meta-C-C6-N unit appears to be particularly relevant in view of the widespread occurrance of this unit and its association with a variety of other building blocks in Streptomyces antibiotics.
| Hornemann, U; Otto, C J; Hoffman, G G et al. (1987) Spectinomycin resistance and associated DNA amplification in Streptomyces achromogenes subsp. rubradiris. J Bacteriol 169:2360-6 |
