Production of polyhydroxyalkanoates with high 3-hydroxydodecanoate monomer content by fadB and fadA knockout mutant of Pseudomonas putida KT2442.
Pseudomonas putida KT2442 produces medium-chain-length (MCL) polyhydroxyalkanoates (PHA) consisting of 3-hydroxyhexanoate (HHx), 3-hydroxyoctanoate (HO), 3-hydroxydecanoate (HD), and 3-hydroxydodecanoate (HDD) from a wide-range of carbon sources. In this study, fadA and fadB genes encoding 3-ketoacyl-CoA thiolase and 3-hydroxyacyl-CoA dehydrogenase in P. putida KT2442 were knocked out to weaken the beta-oxidation pathway. Two-step culture was proven as the optimal method for PHA production in the mutant termed P. putida KTOY06. In a shake-flask culture, when dodecanoate was used as a carbon source, P. putida KTOY06 accumulated 84 wt % PHA, much higher than 50 wt % PHA in its wild type KT2442. The PHA monomer composition was completely different: the HDD fraction in PHA produced by KTOY06 was 41 mol %, much higher compared with 7.5 mol % only in KT2442. The fermentor-scale culture indicated the HDD fraction in PHA decreased during the culture time from 35 to 25 mol % in a one-step fermentation process or from 75 to 49 mol % in a two-step fermentation process. It is for the first time that PHA with a dominant HDD fraction was produced. Thermal and mechanical properties assays indicated that this new type PHA with a high HDD fraction had higher crystallinity and tensile strength than PHA with a low HDD fraction did, demonstrating an improved application property.
Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China.
Shao-Ping
SP
Ouyang
Rong Cong
RC
Luo
Si-Si
SS
Chen
Qian
Q
Liu
Ahleum
A
Chung
Qiong
Q
Wu
Guo-Qiang
GQ
Chen
D001426:Bacterial Proteins / Q000235:genetics; D001709:Biotechnology / Q000379:methods*; D005285:Fermentation; D005798:Genes, Bacterial / Q000235:genetics; D005818:Genetic Engineering; D007850:Lauric Acids / Q000032:analysis / Q000378:metabolism*; D009154:Mutation; D011091:Polyesters / Q000737:chemistry / Q000378:metabolism*; D016958:Pseudomonas putida / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism
D001426:Bacterial Proteins; D007850:Lauric Acids; D011091:Polyesters; C022145:3-hydroxydodecanoic acid