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KMID : 0380020090240030279
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Korean Journal of Biotechnology and Bioengineering
2009 Volume.24 No. 3 p.279 ~ p.286
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Inhibition of Oligomycin Biosynthesis by olmA5 Gene Knock-out in Streptomyces avermitilis
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Kang Hyun-Woo
Ryu Yeon-Woo
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Abstract
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Streptomyces is well known for their ability to synthesize enormous varieties of antibiotics as secondary metabolites. Among them, S. avermitilis produces avermectins, a group of antiparasitic agents used in human and veterinary medicine. However, S. avermitilis also produces oligomycin, which is a potential toxic inhibitor of oxidative phosphorylation in mammalian cells. Therefore, we decided to disrupt oligomycin synthetase gene to prevent co-production of oligomycin in S. avermitilis. To create plasmid for disruption, the smallest gene of oligomycin synthetase gene cluster was obtained by PCR from S. avermitilis chromosome. Then, apramycin resistance gene was inserted in oligomycin synthetase gene for selection. After transformation of this plasmid, oligomycin synthetase gene (olmA5) in the chromosome was displaced with disruption cassette on the plasmid via homologous recombination. As a result of this gene replacement, we obtained mutants (olmA5::apra) that no longer makes the toxic oligomycin. And the mutants confirmed by PCR and HPLC analysis. However, showed no increasement of avermectin production in the mutant was observed.
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KEYWORD
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Streptomyces avermitilis, avermectin, oligomycin, gene knock, homologous recombination
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