Microbial production of 3-hydroxydodecanoic acid by pha operon and fadBA knockout mutant of Pseudomonas putida KT2442 harboring tesB gene.
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.
Chung
Ahleum
A
Beijing Key Laboratory of Protein Therapeutics, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China.
Liu
Qian
Q
Ouyang
Shao-Ping
SP
Wu
Qiong
Q
Chen
Guo-Qiang
GQ
D015094:3-Hydroxyacyl CoA Dehydrogenases / Q000235:genetics / Q000378:metabolism; D000102:Acetyl-CoA C-Acyltransferase / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D064429:Fatty Acid Synthases / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D055786:Gene Knockout Techniques; D007850:Lauric Acids / Q000378:metabolism*; D009154:Mutation; D009876:Operon; D016958:Pseudomonas putida / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D013869:Thiolester Hydrolases / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D007850:Lauric Acids; C022145:3-hydroxydodecanoic acid; D015094:3-Hydroxyacyl CoA Dehydrogenases; D000102:Acetyl-CoA C-Acyltransferase; D064429:Fatty Acid Synthases; D013869:Thiolester Hydrolases; C022738:thioesterase II