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http://dx.doi.org/10.4014/jmb.0907.07020

Enhancement of Clavulanic Acid Production by Expressing Regulatory Genes in gap Gene Deletion Mutant of Streptomyces clavuligerus NRRL3585  

Jnawali, Hum Nath (Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, SunMoon University)
Lee, Hei-Chan (Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, SunMoon University)
Sohng, Jae-Kyung (Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, SunMoon University)
Publication Information
Journal of Microbiology and Biotechnology / v.20, no.1, 2010 , pp. 146-152 More about this Journal
Abstract
Streptomyces clavuligerus NRRL3585 produces a clinically important $\beta$-lactamase inhibitor, clavulanic acid (CA). In order to increase the production of CA, the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene (gap) was deleted in S. clavuligerus NRRL3585 to overcome the limited glyceraldehyde-3-phosphate pool; the replicative and integrative expressions of ccaR (specific regulator of the CA biosynthetic operon) and claR (Lys-type transcriptional activator) genes were transformed together into a deletion mutant to improve clavulanic acid production. We constructed two recombinant plasmids to enhance the production of CA in the gap1 deletion mutant of S. clavuligerus NRRL3585: pHN11 was constructed for overexpression of ccaR-claR, whereas pHN12 was constructed for their chromosomal integration. Both pHN11 and pHN12 transformants enhanced the production of CA by 2.59-fold and 5.85-fold, respectively, compared with the gap1 deletion mutant. For further enhancement of CA, we fed the pHN11 and pHN12 transformants ornithine and glycerol. Compared with the gap1 deletion mutant, ornithine increased CA production by 3.24- and 6.51-fold in the pHN11 and pHN12 transformants, respectively, glycerol increased CA by 2.96- and 6.21-fold, respectively, and ornithine and glycerol together increased CA by 3.72- and 7.02-fold, respectively.
Keywords
Streptomyces clavuligerus NRRL3585; clavulanic acid; regulatory gene; glyceraldehyde-3-phosphate dehydrogenase; fed-batch fermentation; metabolic engineering;
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