Inactivating NADH:Quinone Oxidoreductases Affects The Growth And Metabolism Of Klebsiella Pneumoniae

The intracellular NADH/NAD+ratio is a key fator affecting the metabolism of bacteria.Previous studies for modulating NADH/NAD+ balance to increase the 1,3-propanediol and 2,3-butanediol production of Klebsiella pneumoniae were focused on knocking out NADH-dependent metabolic byproduct pathways or over-expressing NADH regeneration pathways.Few studies have been done to investigate the effects of respiratory chains on those diols production.However respiratory chain is important for the balance of intracellular redox balance.It has been successfully used in the production of anaerobic fermentation products through modulating NADH/NADT balance by respiratory chain in Escherichia coli and Zymomonas mobili.As the electron entry site of bacterial respiratory chain,NADH:quinone oxidoreductases(NQOs)provide an export of intracellular NADH,which could affect the bio-products formation in K.pneumoniae.K.pneumoniae owns three kinds of NQOs,which are NDH-1,NDH-2 and NQR.In this study,inactivating of NQOs,alone or together,on cell metabolism were investigated under different culture conditions.It was found that the growth defect occurred in the NDH-1-NDH-2 double and NDH-1-NDH-2-NQR triple deficient mutants.Based on the experiments,NQR was only effect under aerobic condition,and the growth defect caused by inactivating of NQOs was probably due to the damage of respiratory chain.The experiments data also showed that K.pneumoniae can flexibly use NQOs to maintain the normal growth in single NQO-deficient mutants.Interestingly,under aerobic culture,inactivating of NDH-1 resulted in high intracellular NADH/NAD+ratio,which was proven to be beneficial for 2,3-butanediol production.Compared with the parent strain,the 2,3-butanediol production by NDH-1-deficient mutant increased by 46%and 62%in glyceroland glucose-based mediums respectively.Thus,the result presented here suggested a practicable strategy through metabolic engineering of respiratory chains to promote the biosynthesis of 2,3-butanediol in K.pneumoniae.