[P006] Poster

1:00 PM - 1:50 PM JST(4:00 AM - 4:50 AM UTC)

[P03-384] Analysis of cyanobacterial D-lactate production mechanism using dynamic metabolomics

○Mami Matsuda¹, Tomohisa Hasunuma¹ (1. Kobe University (Japan))

Keywords:

Cyanobacteria,D-Lactate,Metabolomics,Metabolic engineering

D-Lactate is one of the most valuable compounds for manufacturing biobased polymers. Poly(lactic acid) (PLA), which is one of the major biobased polymers composed of lactate, iswidely used as a biodegradable polyester. Stereocomplex PLA is formed by blending enantiomeric PLAs, poly(L-lactide) and poly(D-lactide), and prominently exhibits enhanced mechanical properties as well as heat and hydrolysis resistance. The large-scale industrial production of optically pure L-lactate has been accomplished. On the other hand, biorefineries demand the availability of optically pure D-lactate, an issue that remains to be resolved.Synechocystis sp. PCC6803 (hereafter Synechocystis 6803) accumulates the storage of polysaccharide glycogen from carbon dioxide during oxygenic photosynthesis, and then catabolizes the stored carbohydrate molecule via autofermentation under dark anoxic conditions. In such conditions, Synechocystis 6803 excretes D-lactate. Here, we have investigated the significance of endogenous malic enzyme (ME), which catalyzes the oxidative decarboxylation of malate to pyruvate, in D-lactate biosynthesis in Synechocystis 6803. D-Lactate levels were increased by 2-fold in ME overexpressing strains. To elucidate the alteration in metabolism in Synechocystis 6803, metabolome analysis was performed. In the present study, dynamic metabolomics with ¹³C-bicarbonate revealed that overexpression of ME led to increased turnover rates in malate and pyruvate metabolism; in contrast, deletion of ME resulted in increased pool sizes of glycolytic intermediates, probably due to sequential feedback inhibition, initially triggered by malate accumulation.Finally, both the loss of the acetate kinase gene and overexpression of endogenous D-lactate dehydrogenase, concurrent with ME overexpression, resulted in the highest production of D-lactate (26.6 g/L) with an initial cell concentration of 75 g-dry cell weight/L after 72 h fermentation.