초록
<P><B>ABSTRACT</B></P><P>CcmR, a LysR‐type transcriptional regulator, represses the genes encoding components of the high‐affinity carbon concentration mechanism in cyanobacteria. Unexpectedly, deletion of the <I>ccmR</I> gene was found to alter the expression of the terminal oxidase and fermentative genes, especially the hydrogenase operon in the cyanobacterium <I>Synechococcus sp</I>. PCC 7002. Consistent with the transcriptomic data, the deletion strain exhibits flux increases (30–50%) in both aerobic O<SUB>2</SUB> respiration and anaerobic H<SUB>2</SUB> evolution. To understand how CcmR influences anaerobic metabolism, the kinetics of autofermentation were investigated following photoautotrophic growth. The autofermentative H<SUB>2</SUB> yield increased by 50% in the CcmR deletion strain compared to the wild‐type strain, and increased to 160% (within 20 h) upon continuous removal of H<SUB>2</SUB> from the medium (“milking”) to suppress H<SUB>2</SUB> uptake. Consistent with this greater reductant flux to H<SUB>2</SUB>, the mutant excreted less lactate during autofermentation (NAD(P)H consuming pathway). To enhance the rate of NADH production during anaerobic metabolism, the <I>ccmR</I> mutant was engineered to introduce GAPDH overexpression (more NADH production) and LDH deletion (less NADH consumption). The triple mutant (<I>ccmR</I> deletion + GAPDH overexpression + LDH deletion) showed 6–8‐fold greater H<SUB>2</SUB> yield than the WT strain, achieving conversion rates of 17 nmol 10<SUP>8</SUP> cells<SUP>−1</SUP> h<SUP>−1</SUP> and yield of 0.87 H<SUB>2</SUB> per glucose equivalent (8.9% theoretical maximum). Simultaneous monitoring of the intracellular NAD(P)H concentration and H<SUB>2</SUB> production rate by these mutants reveals an inverse correspondence between these variables indicating hydrogenase‐dependent H<SUB>2</SUB> production as a major sink for consuming NAD(P)H in preference to excretion of reduced carbon as lactate during fermentation. Biotechnol. Bioeng. 2016;113: 1448–1459. © 2015 Wiley Periodicals, Inc.</P>