Enhanced production of poly-3-hydroxybutyrate by Escherichia coli over-expressing multiple copies of NAD kinase integrated in the host genome.
OBJECTIVE
With the help of attB-attP recombination technique, multiple copies of yfjB gene encoding the NAD kinase of Escherichia coli were inserted into the host chromosome to promote NADPH-dependent poly-3-hydroxybutyrate (PHB) production.


RESULTS
The yfjB integration mutant E. coli T2 harbored a similar metabolic profile to that of the wild type control. When PHB biosynthesis operon was introduced, the yfjB integration mutant produced 3 g PHB l(-1) from 18.2 g glucose l(-1), while the wild type consumed 15.7 g glucose l(-1) to afford 2.34 g PHB l(-1) in 48 h of shake-flask cultivation. Transcriptional analysis showed that the transcription levels of genes within the PHB biosynthesis operon were increased by six to eightfold with yfj Bover-expression, which may be the primary reason for the improved PHB production.


CONCLUSIONS
A practical method is demonstrated to construct genetically-stable strains harboring extra copies of NAD kinase to enhance NADPH-dependent reactions.
Beijing Key Lab of Bioprocess, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, China.
Jie
J
Zhang
Xue
X
Gao
Peng-Hui
PH
Hong
Zheng-Jun
ZJ
Li
Tian-Wei
TW
Tan
D053898:Biosynthetic Pathways / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism*; D020869:Gene Expression Profiling; D055879:Gene Knock-In Techniques; D016680:Genome, Bacterial; D005947:Glucose / Q000378:metabolism; D006885:Hydroxybutyrates / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D009249:NADP / Q000378:metabolism; D010770:Phosphotransferases / Q000235:genetics* / Q000378:metabolism*; D011091:Polyesters / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014158:Transcription, Genetic; D000094122:Polyhydroxybutyrates
D029968:Escherichia coli Proteins; D005947:Glucose; D006885:Hydroxybutyrates; D009249:NADP; D010770:Phosphotransferases; D011091:Polyesters; D011994:Recombinant Proteins; C003182:poly-beta-hydroxybutyrate; D000094122:Polyhydroxybutyrates; C434675:NadK protein, E coli