Activity and functional properties of the isocitrate lyase in the cyanobacterium Cyanothece sp. PCC 7424.

Cyanobacteria are ubiquitous photoautotrophs that assimilate atmospheric CO2 as their main source of carbon. Several cyanobacteria are known to be facultative heterotrophs that are able to grow on diverse carbon sources. For selected strains, assimilation of organic acids and mixotrophic growth on acetate has been reported for decades. However, evidence for the existence of a functional glyoxylate shunt in cyanobacteria has long been contradictory and unclear. Genes coding for isocitrate lyase (ICL) and malate synthase were recently identified in two strains of the genus Cyanothece, and the existence of the complete glyoxylate shunt was verified in a strain of Chlorogloeopsis fritschii. Here, we report that the gene PCC7424_4054 of the strain Cyanothece sp. PCC 7424 encodes an enzymatically active protein that catalyses the reaction of ICL, an enzyme that is specific for the glyoxylate shunt. We demonstrate that ICL activity is induced under alternating day/night cycles and acetate-supplemented cultures exhibit enhanced growth. In contrast, growth under constant light did not result in any detectable ICL activity or enhanced growth of acetate-supplemented cultures. Furthermore, our results indicate that, despite the presence of a glyoxylate shunt, acetate does not support continued heterotrophic growth and cell proliferation. The functional validation of the ICL is supplemented with a bioinformatics analysis of enzymes that co-occur with the glyoxylate shunt. We hypothesize that the glyoxylate shunt in Cyanothece sp. PCC 7424, and possibly other nitrogen-fixing cyanobacteria, is an adaptation to a specific ecological niche and supports assimilation of nitrogen or organic compounds during the night phase.

Flux balance analysis of cyanobacterial metabolism: the metabolic network of Synechocystis sp. PCC 6803.

Cyanobacteria are versatile unicellular phototrophic microorganisms that are highly abundant in many environments. Owing to their capability to utilize solar energy and atmospheric carbon dioxide for growth, cyanobacteria are increasingly recognized as a prolific resource for the synthesis of valuable chemicals and various biofuels. To fully harness the metabolic capabilities of cyanobacteria necessitates an in-depth understanding of the metabolic interconversions taking place during phototrophic growth, as provided by genome-scale reconstructions of microbial organisms. Here we present an extended reconstruction and analysis of the metabolic network of the unicellular cyanobacterium Synechocystis sp. PCC 6803. Building upon several recent reconstructions of cyanobacterial metabolism, unclear reaction steps are experimentally validated and the functional consequences of unknown or dissenting pathway topologies are discussed. The updated model integrates novel results with respect to the cyanobacterial TCA cycle, an alleged glyoxylate shunt, and the role of photorespiration in cellular growth. Going beyond conventional flux-balance analysis, we extend the computational analysis to diurnal light/dark cycles of cyanobacterial metabolism.

Impaired glycogen synthesis causes metabolic overflow reactions and affects stress responses in the cyanobacterium Synechocystis sp. PCC 6803.

The biosynthesis of glycogen or starch is one of the main strategies developed by living organisms for the intracellular storage of carbon and energy. In phototrophic organisms, such polyglucans accumulate due to carbon fixation during photosynthesis and are used to provide maintenance energy for cell integrity, function and viability in dark periods. Moreover, it is assumed that glycogen enables cyanobacteria to cope with transient starvation conditions, as observed in most micro-organisms. Here, glycogen accumulates when an appropriate carbon source is available in sufficient amounts but growth is inhibited by lack of other nutrients. In this study, the role of glycogen in energy and carbon metabolism of phototrophic cyanobacteria was first analysed via a comparative physiological and metabolic characterization of knockout mutants defective in glycogen synthesis. We first proved the role of glycogen as a respiratory substrate in periods of darkness, the role of glycogen as a reserve to survive starvation periods such as nitrogen depletion and the role of glycogen synthesis as an ameliorator of carbon excess conditions in the model organism Synechocystis sp. PCC 6803. We provide striking new insights into the complex carbon and nitrogen metabolism of non-diazotrophic cyanobacteria: a phenotype of sensitivity to photomixotrophic conditions and of reduced glucose uptake, a non-bleaching phenotype based on an impaired acclimation response to nitrogen depletion and furthermore a phenotype of energy spilling. This study shows that the analysis of deficiencies in glycogen metabolism is a valuable tool for the identification of metabolic regulatory principles and signals.