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Background  2,3-Butanediol (2,3-BD) is a high-value chemical usually produced petrochemically but which can also be synthesized by some bacteria. To date,  Klebsiella pneumoniae  is the most powerful 2,3-BD producer which can utilize a wide range of substrates. However, many by-products are also produced by  K. pneumoniae , such as ethanol, lactate, and acetate, which negatively regulate the 2,3-BD yield and increase the costs of downstream separation and purification.    Results  In this study, we constructed  K. pneumoniae  mutants with lactate dehydrogenase (LDH), acetaldehyde dehydrogenase (ADH), and phosphotransacetylase (PTA) deletion individually by suicide vector conjugation.  These mutants showed different behavior of production formation. Knock out of  ldhA  had little influence on the yield of 2,3-BD, whereas knock out of  adhE  or  pta  significantly improved the formation of 2,3-BD. The accumulation of the intermediate of 2,3-BD biosynthesis, acetoin, was decreased in all the mutants. The mutants were then tested in five different carbon sources and increased 2,3-BD was observed. Also a double mutant strain with deletion of  adhE  and  ldhA  was constructed which resulted in accelerated fermentation and higher 2,3-BD production. In fed-batch culture this strain achieved more than 100 g/L 2,3-BD from glucose with a relatively high yield of 0.49 g/g.    Conclusion  2,3-BD production was dramatically improved with the inactivation of  adhE  and  pta.  The inactivation of  ldhA  could advance faster cell growth and shorter fermentation time. The double mutant strain with deletion of  adhE  and  ldhA  resulted in accelerated fermentation and higher 2,3-BD production. These results provide new insights for industrial production of 2,3-BD by  K. pneumoniae .

biotechnology for biofuels

Effect of the inactivation of lactate dehydrogenase, ethanol dehydrogenase, and phosphotransacetylase on 2,3-butanediol production in Klebsiella pneumoniae strain