The Study On The Pathway Of Pyruvate Metabolism In Klebsiella Pneumoniae

Pyruvate is an important metabolic node,and metabolic flux from pyruvate to acetyl-CoA is concentrated in Escherichia coli.It is generally concluded that pyruvate dehydrogenase(PDHC)is responsible for catalyzing pyruvate to acetyl-CoA under aerobic growth condition,while pyruvate formate lyase(PFL)plays for catalyzing pyruvate to acetyl-CoA under anaerobic fermentation.Howerer,in recent years,it has been reported that PDHC also works under anaerobic condition.Till to now,there are few studies focus on the pyruvate-acetyl-CoA metabolism in Klebsiella pneumonia.Thus,it is important to study the mechanism of carbon flow in pyruvate node of K.pneumoniae,which is a high-yielding 1,3-propanediol-producing microorganism.In this study,PDHC-deficient K.pneumoniae,strain of KG-PD and PFL-deficient K.pneumoniae,strain of KG-PF were obtained by knocking out the related genes through the Red homologous recombination technique.The growth and metabolism of PDHC-or PFL-deficient mutant in glycerol-based medium were analyzed and compared with those of the parent strain.The results showed that the cell growth of KG-PD in the aerobic and anaerobic cultures were seriously inhibited,and the inhibition of cell growth under aerobic condition was more serious.So,combined the emzyme activites of PDHC,it can be,deduced that,under aerobic condition,pyruvate catalyzed to acetyl-CoA is mainly dependent on PDHC in K.pneumoniae.Howerer,under anaerobic condition,the PDHC activity was only 50%of that of PFL,blocking of PDHC also resulted in more growth defect than blocking of PFL.Subsequently,combined with the requirement of CO2 and intracellular redox status,it was presumed that the critical role of PDHC was to provide NADH for the anaerobic growth of K.pneumoniae.This presumption was confirmed in the PDHC-deficient mutant by further blocking one of the formate dehydrogenases,FdnGHI.Based on our data,the yield of 1,3-propanediol was improved in mutant strain of KG-PF under both aerobic and anaerobic conditions,and strain KG-PF could be used as a potential dominant strain for high 1,3-propanediol production and,the strain of KG-PDN was showed an excellent growth and high acetoin production ability under aerobic culture.