Targeted disruption of homoserine dehydrogenase gene in Streptomyces clavuligerus and its effects on cephamycin C production

Çaydaşı (Koca), Ayşe
The members of the genus Streptomyces are well-known for their capacity to synthesize a vast repertoire of secondary metabolites, including many useful antibiotics and proteins. Streptomyces clavuligerus is the producer of the medically important β-lactam antibiotics such as cephamycin C and the potent β-lactamase inhibitor clavulanic acid. The aspartate pathway of S. clavuligerus is an important primary metabolic pathway providing substrates for β-lactam synthesis. This pathway uses L-aspartic acid as the precursor for the biosynthesis of the amino acids L-lysine, L-methionine, L-isoleucine, L-threonine and several important metabolic intermediates. L-α-aminoadipic acid (α-AAA) required for β-lactam synthesis is a catabolic product of L-lysine produced from the lysine branch of the aspartate pathway. The carbon flow through the L-lysine-specific branch of aspartate pathway is limiting for the formation of cephamycin C. Formation of L-homoserine from aspartate semialdehyde (ASA) is the first step of the other branch of the aspartate pathway leading to L-threonine, L-isoleucine and L-methionine synthesis and is catalyzed by homoserine dehydrogenase (HSD, EC Regulation of the activity or biosynthesis of the HSD of S. clavuligerus determines the availability of ASA for the biosynthesis of L-lysine and α-AAA. The gene encoding for homoserine dehydrogenase (hom) was previously cloned from S. clavuligerus NRRL 3585 and characterized in our laboratory. In this study, the hom gene was disrupted via insertion of a kanamycin resistance cassette into this gene which was subsequently transferred to S. clavuligerus cells using the Streptomyces plasmid vector pIJ486. A hom mutant of S. clavuligerus (AK39) was formed through integration into the chromosome by double crossing over and the effects of hom disruption on cephamycin C yields were investigated. Disruption of hom gene resulted in a 1.7 to 2.0 fold increase in specific cephamycin C production in chemically defined medium (CDM).


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Citation Formats
A. Çaydaşı (Koca), “Targeted disruption of homoserine dehydrogenase gene in Streptomyces clavuligerus and its effects on cephamycin C production,” M.S. - Master of Science, Middle East Technical University, 2006.