Dramatic Activation of Antibiotic Production in Streptomyces coelicolor by Cumulative Drug-Resistance Mutations

National Food Research Institute, Tsukuba, Ibaraki 305-8642, Japan

^* To whom correspondence should be addressed. Email: kochi{at}affrc.go.jp.

	Abstract

We recently described a new method to activate antibiotic production in bacteria, by introducing a mutation conferring resistance to a drug, such as streptomycin, rifampicin, paromomycin, or gentamicin. This method, however, enhanced antibiotic production only by up to an order of magnitude. Working with Streptomyces coelicolor A3(2), we established a method for dramatic activation of antibiotic production by sequential introduction of multiple drug-resistance mutations. Septuple and octuple mutants, C7 and C8, thus obtained by screening for resistance to seven or eight drugs produced huge amounts (1.63 g/l) of the polyketide antibiotic actinorhodin, 180-fold higher than produced by wild-type. This dramatic overproduction was due to the acquisition of mutant ribosomes, with aberrant protein and ppGpp synthesis activity, as demonstrated by in vitro protein synthesis assay and by abolition of antibiotic overproduction with relA disruption. This new approach, called "ribosome engineering", requires less time, cost, and labor than other methods, and may be widely utilized for bacterial strain improvement.