| 초록 |
The glycolitic metabolism of A. eutrophus was investigated using Poly-β-hydroxybutyrate (PHB) negative mutant and transformants harboring cloned phbC and phbCAB genes to reinforce the PHB biosynthesis enzymes. The PHB negative mutant strain was absolutly defected in PHB synthase, meanwhile, partially active in β-ketothiolase and acetoacetyl-CoA reductase. The mutant strain excreted substantial amount of intermediates of glycolitic metabolism at the late growth stage, expecially pyruvate. The excretion of pyruvate in mutant strain was due to the inhibition of pyruvate dehydrogenase caused by the accumulation of acetyl-CoA. The transformant harboring cloned phbC gene, reinforced PHB synthase, reduced the excretion of pyruvate mostly, and the transformant reinforced cloned phbCAB gene containing all three enzymes did not excrete pyruvate. By increasing activities of β-ketothiolase and acetoacetyl-CoA reductase, the rate of PHB biosynthesis was accelerated, meanwhile, the excretion of pyruvate was decresed due to the reduced accumulation of acetyl-CoA. The β-ketothiolase was found to be the most critical enzyme related to glycolitic metabolism especially for utilization of pyruvate hence determine the rate of PHB biosynthesis in A. eutrophus.
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