Expression of the pfl Gene and Resulting Metabolite Flux Distribution in nuo and ackA-pta E.coli Mutant Strains

摘要

Our laboratory previously studied the interaction between nuo and the acetate-producing pathway encoded by ackA-pta in Escherichia coli.We examined metabolic patterns,particularly the ethanol and acetate production rates,of several mutant strains grown under anaerobic growth conditions.Since the pyruvate formate-lyase (PFL)pathway is the major route for acetyl-CoA and formate production under anaerobic conditions,we examined the effects of nuo and ackA/pta mutations on the expression of pyruvate formate-lyase (pfl)under anaerobic conditions.The ackA-pta mutant has a pfl::lacZ expression level much higher than that of the wild-type strain,and cultures also exhibit the highest ethanol production.Real-time PCR demonstrated that the adhE gene expression in the ack-pta mutant strain was approximately 100 fold that of the same gene in the ackA-pta nuo mutant strain.This result correlates with the observed ethanol production rates in cultures of the strain.However,the lack of exact correlation between the ethanol production rates and the RT-PCR data suggests additional regulation actions at the posttranslation level.In addition,the activity of the pfl gene as indicated by mRNA levels was also considerably greater in the ack-pta mutant.We can conclude that deletions of nuo and ack/pta can partially affect the expression of the genes encoding adhE and pfl under anaerobic conditions.