Dataset Information

Over-expression of an electron transport protein OmcS provides sufficient NADH for D-lactate production in cyanobacterium.

ABSTRACT:

Background

An efficient supply of reducing equivalent is essential for chemicals production by engineered microbes. In phototrophic microbes, the NADPH generated from photosynthesis is the dominant form of reducing equivalent. However, most dehydrogenases prefer to utilize NADH as a cofactor. Thus, sufficient NADH supply is crucial to produce dehydrogenase-derived chemicals in cyanobacteria. Photosynthetic electron is the sole energy source and excess electrons are wasted in the light reactions of photosynthesis.

Results

Here we propose a novel strategy to direct the electrons to generate more ATP from light reactions to provide sufficient NADH for lactate production. To this end, we introduced an electron transport protein-encoding gene omcS into cyanobacterium Synechococcus elongatus UTEX 2973 and demonstrated that the introduced OmcS directs excess electrons from plastoquinone (PQ) to photosystem I (PSI) to stimulate cyclic electron transfer (CET). As a result, an approximately 30% increased intracellular ATP, 60% increased intracellular NADH concentrations and up to 60% increased biomass production with fourfold increased D-lactate production were achieved. Comparative transcriptome analysis showed upregulation of proteins involved in linear electron transfer (LET), CET, and downregulation of proteins involved in respiratory electron transfer (RET), giving hints to understand the increased levels of ATP and NADH.

Conclusions

This strategy provides a novel orthologous way to improve photosynthesis via enhancing CET and supply sufficient NADH for the photosynthetic production of chemicals.

SUBMITTER: Meng H

PROVIDER: S-EPMC8082822 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:In order to obtain a global view of energy metabolism pathways of the sulfate-reducer Desulfovibrio vulgaris Hildenborough and the proteins involved therein whole-genome microarrays were used to compare the transcriptional response of cells grown with hydrogen/sulfate, pyruvate/sulfate, lactate/thiosulfate, and pyruvate with limiting sulfate, relative to growth in standard lactate/sulfate condition. Growth with hydrogen/sulfate showed the largest number of differently expressed genes and the largest changes in expression levels. The most up-regulated energy metabolism genes were those coding for the periplasmic [NiFeSe] hydrogenase, followed by the Ech hydrogenase, and the most down-regulated were genes coding for the Coo hydrogenase. The results point to the involvement of formate cycling and the ethanol pathway during growth on hydrogen, whereas there is evidence for CO cycling during growth on lactate and pyruvate, but not on H2. Growth with thiosulfate showed the hallmarks of a reduced energy status of the cells with down regulation of the ATP synthase and the Qmo and Dsr complexes., whereas growth with pyruvate showed the smallest differences but an increased role for the Ech hydrogenase.that in this case functions in reverse from the case of growth with H2. The multiple periplasmic hydrogenases and formate dehydrogenases, do not display the same regulation pattern showing that their metabolic roles are not totally interchangeable. This result together with the observation that several genes coding for proteins that have not been biochemically characterised were considerably affected in this study, reveals a more complex energy metabolism than previously considered and provides guidance for further studies. Keywords: Growth protocol Desulfovibrio vulgaris from our laboratory culture collection was cultured at 37°C with hydrogen as the only electron donor (with sulfate as the electron acceptor) to mid-log phase. Cultures were also cultivated similarly using lactate as the sole electron donor to mid-log phase. Gene expression profiles of cultures grown with hydrogen as the electron donor were compared with those of the cultures grown with lactate as the electron donor for sulfate reduction. Total RNA was harvested from four replicate cultures for microarray analysis. RNA extraction, purification, and labeling were performed independently on each cell sample.Two samples of each total RNA preparation were labeled, one with Cy3-dUTP and another with Cy5-dUTP for microarray hybridization (dye swap).