TY - JOUR AU - Matsumoto, Ken’ichiro AU - Tobitani, Kota AU - Aoki, Shunsuke AU - Song, Yuyang AU - Ooi, Toshihiko AU - Taguchi, Seiichi PY - 2014 DA - 2014/11/18 TI - Improved production of poly(lactic acid)-like polyester based on metabolite analysis to address the rate-limiting step JO - AMB Express SP - 83 VL - 4 IS - 1 AB - The biosynthesis of poly(lactic acid) (PLA)-like polymers, composed of >99 mol% lactate and a trace amount of 3-hydroxybutyrate, in engineered Corynebacterium glutamicum consists of two steps; the generation of the monomer substrate lactyl-coenzyme A (CoA) and the polyhydroxyalkanoate (PHA) synthase-catalyzed polymerization of lactyl-CoA. In order to increase polymer productivity, we explored the rate-limiting step in PLA-like polymer synthesis based on quantitative metabolite analysis using liquid chromatography mass spectroscopy (LC-MS). A significant pool of lactyl-CoA was found during polymer synthesis. This result suggested that the rate-limitation occurred at the polymerization step. Accordingly, the expression level of PHA synthase was increased by means of codon-optimization of the corresponding gene that consequently led to an increase in polymer content by 4.4-fold compared to the control. Notably, the codon-optimization did not significantly affect the concentration of lactyl-CoA, suggesting that the polymerization reaction was still the rate-limiting step upon the overexpression of PHA synthase. Another important finding was that the generation of lactyl-CoA was concomitant with a decrease in the acetyl-CoA level, indicating that acetyl-CoA served as a CoA donor for lactyl-CoA synthesis. These results show that obtaining information on the metabolite concentrations is highly useful for improving PLA-like polymer production. This strategy should be applicable to a wide range of PHA-producing systems. SN - 2191-0855 UR - https://doi.org/10.1186/s13568-014-0083-2 DO - 10.1186/s13568-014-0083-2 ID - Matsumoto2014 ER -