Journal
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
Volume 83, Issue 3, Pages 513-519Publisher
SPRINGER
DOI: 10.1007/s00253-009-1919-6
Keywords
Polyhydroxyalkanoates; PHA; 3-hydroxyalkanoic acid; 3HA; 3-hydroxydodecanoic acid; 3HDD; Pseudomonas putida; 3-Ketoacyl-CoA thiolase; 3-Hydroxyacyl-CoA dehydrogenase; Thioesterase II; TesB; fadBA
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Funding
- National High Tech 863 [2006AA02Z242, 2006AA020104]
- State Basic Science Foundation 973 [2007CB707804]
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To produce extracellular chiral 3-hydroxyacyl acids (3HA) by fermentation, a novel pathway was constructed by expressing tesB gene encoding thioesterase II into Pseudomonas putida KTOY01, which was a polyhydroxyalkanoate (PHA) synthesis operon knockout mutant. 3HA mixtures of 0.35 g/l consisting of 3-hydroxyhexanoate, 3-hydroxyoctanoate, 3-hydroxydecanoate, and 3-hydroxydodecanoate (3HDD) were produced in shake-flask study using dodecanoate as a sole carbon source. Additional knockout of fadB and fadA genes encoding 3-ketoacyl-CoA thiolase and 3-hydroxyacyl-CoA dehydrogenase in P. putida KTOY01 led to the weakening of the beta-oxidation pathway. The fadBA and PHA synthesis operon knockout mutant P. putida KTOY07 expressing tesB gene produced 2.44 g/l 3HA, significantly more than that of the beta-oxidation intact mutant. The 3HA mixture contained 90 mol% 3HDD as a dominant component. A fed-batch fermentation process carried out in a 6-l automatic fermentor produced 7.27 g/l extracellular 3HA containing 96 mol% fraction of 3HDD after 28 h of growth. For the first time, it became possible to produce 3HDD-dominant 3HA monomers.
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