Monday, August 2, 2010
Pacific Concourse (Hyatt Regency San Francisco)
Streptomyces clavuligerus, a filamentous bacterium has remarkable capabilities to produce 21 secondary metabolites of diverse chemical structures and biological activities. Clavulanic acid (C.A.) is an economically valuable product and is an important β-lactamase inhibitor. C.A. inhibits choromosal and plasmid β-lactamase produced by Gram-positive and Gram-negative bacteria. Mutagenesis is a powerful method that has been used for enhancing of C.A. production by S. clavuligerus. However, there is no report on mutagenesis by N-methyl-N-nitro-N-nitrosoguanidine (MNNG). In this study, MNNG was used to improve the level of C.A. production by S. clavuligerus. Spore suspension of S. clavuligerus DSM 738 was exposed to various doses MNNG at different pHs. The survivors were isolated by spreading the spore suspension on ISP2 agar plates. The plates were incubated in 28 ºC for 14 days. Overproducing mutants were isolated by bioassay method against Klebsiella pneumoniea in the medium containing 7 µg/ml penicillin G. Also, concentration of C.A. was measured by HPLC using C-18 column. In 10 mutants, concentration of C.A. was higher than that of the parent strain (S. clavuligerus DSM 738). C.A. concentration in the mutants N31, N47, N28 was 1.6, 1.9 and 2.2 times more than that of the parent strain. The yields of C.A. to biomass (YC.A/Biomass) in N31, N47, N28 and the parent strain were 1.76, 2.14, 2.41, and 1.08, respectively. Hereby we suggest that some genes involved in metabolic pathway of C.A. have been mutated by exposure to MNNG, and these mutants could be useful in industrial production of C.A.