Metabolic engineering of Aeromonas hydrophila for the enhanced production of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate).
Wild-type Aeromonas hydrophila 4AK4 produced 35-45 wt.% poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) consisting of 10-15 mol% 3-hydroxyhexanoate (3HHx). To enhance PHBHHx production, vgb gene encoding Vitreoscilla haemoglobin or fadD gene encoding Escherichia coli acyl-CoA synthase was co-expressed with polyhydroxyalkanoates (PHA) synthesis-related genes including phbAB from Wautersia eutropha and phaPCJ from A. hydrophila. Expression of vgb increased PHBHHx content from 46 to 53 wt.% without affecting the polymer monomers composition, whereas fadD increased both PHBHHx content from 46 to 64 wt.% and its 3HHx fraction from 15 to 24 mol%. Co-expression of vgb or fadD gene with PHA-synthesis-related genes generally increased PHBHHx content over 60 wt.%. Co-expression of phbAB with vgb increased PHBHHx content and concentration up to about 70 wt.% and 4.0 g l-1, respectively. Fermentor study also showed that in the recombinants harboring vgb, CDW, PHBHHx concentration and productivity were significantly elevated up to 54 g l-1, 28.5 g l-1 and 0.791 g l-1 h-1, respectively, suggesting that vgb could promote PHA synthesis. In this strain, lac promoter could be used to constitutively express foreign genes such as phbA and phbB encoding beta-ketothiolase and NADPH-dependent acetoacetyl-CoA reductase of W. eutropha, respectively, without use of IPTG. The results showed that combined expression of different genes was a successful strategy to enhance PHA production, which could be useful for strain development to construct other recombinant PHA-producing strains.
Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing, 100084, China.
Yuan-Zheng
YZ
Qiu
Jing
J
Han
Guo-Qiang
GQ
Chen
D020155:3-Hydroxybutyric Acid / Q000737:chemistry / Q000378:metabolism*; D000102:Acetyl-CoA C-Acyltransferase / Q000235:genetics; D016980:Aeromonas hydrophila / Q000235:genetics* / Q000378:metabolism*; D000445:Aldehyde Oxidoreductases / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D002208:Caproates / Q000737:chemistry / Q000378:metabolism*; D003001:Cloning, Molecular; D003066:Coenzyme A Ligases; D020118:Cupriavidus necator / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D005285:Fermentation; D015870:Gene Expression; D006420:Hemeproteins / Q000235:genetics; D007763:Lac Operon / Q000235:genetics; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011995:Recombination, Genetic; D020589:Vitreoscilla / Q000235:genetics; D000094122:Polyhydroxybutyrates
D020155:3-Hydroxybutyric Acid; D000102:Acetyl-CoA C-Acyltransferase; D000445:Aldehyde Oxidoreductases; D001426:Bacterial Proteins; D002208:Caproates; D003066:Coenzyme A Ligases; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D006420:Hemeproteins; C118828:PHAP protein, Bacteria; C100771:Vgb protein, Bacteria; C115940:poly(3-hydroxybutyrate-co-3-hydroxyhexanoate); D000094122:Polyhydroxybutyrates; C027014:acetaldehyde dehydrogenase (acylating); C085934:acyl-coenzyme A synthetase, E coli