Microbial production of medium-chain-length 3-hydroxyalkanoic acids by recombinant Pseudomonas putida KT2442 harboring genes fadL, fadD and phaZ.
Monomers of microbial polyhydroxyalkanoates, mainly 3-hydroxyhexanoic acid (3HHx) and 3-hydroxyoctanoic acid (3HO), were produced by overexpressing polyhydroxyalkanoates depolymerase gene phaZ, together with putative long-chain fatty acid transport protein fadL of Pseudomonas putida KT2442 and acyl-CoA synthetase (fadD) of Escherichia coli MG1655 in P. putida KT2442. FadL(Pp), which is responsible for free fatty acid transportation from the extracellular environment to the cytoplasm, and FadD(Ec), which activates fatty acid to acyl-CoA, jointly reinforce the fatty acid beta-oxidation pathway. Pseudomonas putida KT2442 (pYZPst01) harboring polyhydroxyalkanoates depolymerase gene phaZ of Pseudomonas stutzeri 1317 produced 1.37 g L(-1) extracellular 3HHx and 3HO in shake flask studies after 48 h in the presence of sodium octanoate as a sole carbon source, while P. putida KT2442 (pYZPst06) harboring phaZ(Pst), fadD(Ec) and fadL(Pp) achieved 2.32 g L(-1) extracellular 3HHx and 3HO monomer production under the same conditions. In a 48-h fed-batch fermentation process conducted in a 6-L fermentor with 3 L sodium octanoate mineral medium, 5.8 g L(-1) extracellular 3HHx and 3HO were obtained in the fermentation broth. This is the first time that medium-chain-length 3-hydroxyalkanoic acids (mcl-3HA) were produced using fadL(Pp) and fadD(Ec) genes combined with the polyhydroxyalkanoates depolymerase gene phaZ.
Yuan
Mei-Qing
MQ
College of Biology Science, China Agricultural University, Beijing, China.
Shi
Zheng-Yu
ZY
Wei
Xiao-Xing
XX
Wu
Qiong
Q
Chen
San-Feng
SF
Chen
Guo-Qiang
GQ
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D019149:Bioreactors; D002208:Caproates / Q000378:metabolism; D002210:Caprylates / Q000378:metabolism; D002265:Carboxylic Ester Hydrolases / Q000235:genetics / Q000378:metabolism*; D003066:Coenzyme A Ligases / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D050612:Fatty Acid Transport Proteins / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D054813:Polyhydroxyalkanoates / Q000096:biosynthesis*; D016958:Pseudomonas putida / Q000235:genetics* / Q000378:metabolism*; D044223:Pseudomonas stutzeri / Q000201:enzymology / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D001426:Bacterial Proteins; D002208:Caproates; D002210:Caprylates; D029968:Escherichia coli Proteins; D050612:Fatty Acid Transport Proteins; D054813:Polyhydroxyalkanoates; D011994:Recombinant Proteins; C045051:3-hydroxyhexanoic acid; C101940:3-hydroxyoctanoic acid; D002265:Carboxylic Ester Hydrolases; C077226:poly(3-hydroxyalkanoic acid) depolymerase; D003066:Coenzyme A Ligases; C085934:acyl-coenzyme A synthetase, E coli; C031492:octanoic acid