An insight into the role of phosphotransacetylase (pta) and the acetate/acetyl-CoA node in Escherichia coli.
BACKGROUND
Acetate metabolism in Escherichia coli plays an important role in the control of the central metabolism and in bioprocess performance. The main problems related to the use of E. coli as cellular factory are i) the deficient utilization of carbon source due to the excretion of acetate during aerobic growth, ii) the inhibition of cellular growth and protein production by acetate and iii) the need for cofactor recycling (namely redox coenzymes and free CoASH) to sustain balanced growth and cellular homeostasis.


RESULTS
This work analyzes the effect of mutations in the acetate excretion/assimilation pathways, acetyl-CoA synthethase (acs) and phosphotransacetylase (pta), in E. coli BW25113 grown on glucose or acetate minimal media. Biomass and metabolite production, redox (NADH/NAD+) and energy (ATP) state, enzyme activities and gene expression profiles related to the central metabolism were analyzed. The knock-out of pta led to a more altered phenotype than that of acs. Deletion of pta reduced the ability to grow on acetate as carbon source and strongly affected the expression of several genes related to central metabolic pathways.


CONCLUSIONS
Results showed that pta limits biomass yield in aerobic glucose cultures, due to acetate production (overflow metabolism) and its inefficient use during glucose starvation. Deletion of pta severely impaired growth on acetate minimal medium and under anaerobiosis due to decreased acetyl-coenzyme A synthethase, glyoxylate shunt and gluconeogenic activities, leading to lower growth rate. When acetate is used as carbon source, the joint expression of pta and acs is crucial for growth and substrate assimilation, while pta deletion severely impaired anaerobic growth. Finally, at an adaptive level, pta deficiency makes the strain more sensitive to environmental changes and de-regulates the central metabolism.
Department of Biochemistry and Molecular Biology B and Immunology, Campus de Espinardo, Universidad de Murcia, E-30100, Spain. saracastano@um.es
Sara
S
Castaño-Cerezo
José M
JM
Pastor
Sergio
S
Renilla
Vicente
V
Bernal
José L
JL
Iborra
Manuel
M
Cánovas
D000085:Acetates / Q000378:metabolism*; D000105:Acetyl Coenzyme A / Q000378:metabolism / Q000502:physiology*; D018533:Biomass; D003066:Coenzyme A Ligases / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D004926:Escherichia coli / Q000201:enzymology* / Q000254:growth & development; D020869:Gene Expression Profiling; D055786:Gene Knockout Techniques; D009154:Mutation; D009243:NAD / Q000378:metabolism; D010709:Phosphate Acetyltransferase / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
D000085:Acetates; D009243:NAD; D000105:Acetyl Coenzyme A; D010709:Phosphate Acetyltransferase; D003066:Coenzyme A Ligases; C101958:acetate-CoA ligase (ADP-forming)