Metabolic Engineering of Klebsiella Pneumoniae for 1-Butanol Production By Using Crude Glycerol

Miaomiao Wamg, Tianwei Tan*

Beijing Key Laboratory of Bioprocess, College of Life Science and Technology

Beijing University of Chemical Technology, Beijing 100029, China

   Seeking alternative energy sources has gained much popularity due to the shortage of fuel and increasing global environmental concerns. During the past decades,ethanol has be seen the major renewable substitute of fuel. However, ethanol is not the ideal one, because of its lower energy density and hygroscopicity-resulting storage and distribution problems. Biobutanol, produced by using various renewable resources, is considered one kind of better alternative fuel. It is able to be synthesized by the engineered Klebsiella pneumoniae using crude glycerol from the production of biodiesel as the sole source. Comparing the relative pathways published on KEGG between Cloustridium acetobutylicum and K. pneumonia, we found that in K. pneumonia there exist some enzymes which have similar catalytical roles with some essential genes needed in butanol synthesis in C. acetobutylicum. Only additional bcd/etfAB, bdhB and bdhA, absent in K. pneumonia, are to be introduced. When constructing the engineered strain, the gene ter, instead of the bcd/etfAB, was introduced with the other two genes. The engineered K. pneumonia has the ability of yielding 18 mg/L butanol using glycerol as the only carbon source by flask fermentation while the native strain produces no butanol. Moreover, the antisense RNA strategy was adopted to reduce the level of relative genes’ mRNA to interfere the synthesis of 1, 3-propanediol and 2, 3-butanediol that were native products during the fermentation. The resultant yield of 1, 3-propanediol was reduced by 90%, 2, 3-butanediol was by 10%, compared to those of the K. pneumonia control strain.