P127: Enantioselective production of 2R,3R-butanediol using recombinant Klebsiella oxytoca

Paenibacillus polymyxa can produce optically active 2R,3R-butanediol (2R,3R-BDO) from glucose however, high titer has not been achieved either by native or recombinant strains. On the other hand Klebsiella oxytoca produces meso-2R,3S-BDO at high titer which is optically inactive. In general, both the forms of BDO are produced from a common intermediate, 2R-acetoin. The present study aimed at the production of optically active 2R,3R-BDO by engineering K. oxytoca. To do so, the budC (acetoin reductase) of K. oxytoca (Ko-budC) was precisely replaced by P. polymyxa budC (Pp-budC). When subjected for a batch fermentation, K. oxytoca_ΔldhA_ΔbudC::PP-budC produced about 90% pure 2R,3R-BDO. The reduced purity was due to the co-production of meso- and 2S,3S-BDO. When diacetyl reducase (dar) which shares high homology with budC was deleted from the same strain, it has produced with the purity of 97% 2R,3R-BDO. Furthermore, a fed-batch fermentation was carried out by feeding glucose intermittently and the strain K. oxytoca_ΔldhA_Δdar_ΔbudC::PP-budC produced 98 g/L of 2R,3R-BDO with the enantiomeric excess of 96%. The overall productivity was 2.04 g/L/h. The enantiopure production of 2R,3R-BDO by recombinant K. oxytoca is reported here for the first time at high titer and productivity.