Synechocystis sp. PCC 6803 Requires the Bidirectional Hydrogenase to Metabolize Glucose and Arginine Under Oxic Conditions

Abstract

The cyanobacterium Synechocystis sp.PCC 6803 possesses a bidirectional NiFe-hydrogenase, HoxEFUYH. It functions to produce hydrogen under dark, fermentative conditions and photoproduces hydrogen when dark-adapted cells are illuminated. Unexpectedly, we found that the deletion of the large subunit of the hydrogenase (HoxH) in Synechocystis leads to an inability to grow on arginine and glucose under continuous light in the presence of oxygen. This is surprising, as the hydrogenase is an oxygen-sensitive enzyme. In wild-type (WT) cells, thylakoid membranes largely disappeared, cyanophycin accumulated, and the plastoquinone (PQ) pool was highly reduced, whereas ΔhoxH cells entered a dormant-like state and neither consumed glucose nor arginine at comparable rates to the WT. Hydrogen production was not traceable in the WT under these conditions. We tested and could show that the hydrogenase does not work as an oxidase on arginine and glucose but has an impact on the redox states of photosynthetic complexes in the presence of oxygen. It acts as an electron valve as an immediate response to the supply of arginine and glucose but supports the input of electrons from arginine and glucose oxidation into the photosynthetic electron chain in the long run, possibly via the NDH-1 complex. Despite the data presented in this study, the latter scenario requires further proof. The exact role of the hydrogenase in the presence of arginine and glucose remains unresolved. In addition, a unique feature of the hydrogenase is its ability to shift electrons between NAD(H), NADP(H), ferredoxin, and flavodoxin, which was recently shown in vitro and might be required for fine-tuning. Taken together, our data show that Synechocystis depends on the hydrogenase to metabolize organic carbon and nitrogen in the presence of oxygen, which might be an explanation for its prevalence in aerobic cyanobacteria.

Arginine inhibits the arginine biosynthesis rate-limiting enzyme and leads to the accumulation of intracellular aspartate in Synechocystis sp. PCC 6803

“…6 b). Synechocystis 6803 produces and accumulates cyanophycin, resulting in increased intracellular arginine levels in BG-11 medium supplemented with 5 mM arginine and 10 mM glucose (Burgstaller et al 2022 ). In this study, the intracellular arginine of these strains with 5 mM arginine was higher than those with 5 mM NaNO 3 (Fig.…”

“…Cyanophycin contains equal molecules of arginine and aspartate, therefore the concentration of cyanophycin equals the amount of arginine. We performed cyanophycin extraction and arginine quantification using the Sakaguchi reaction according to the studies (Burgstaller et al 2022, Messineo 1966) with some modifications. After 3 days of cultivation, Synechocystis 6803 cells [OD 730 × culture volume (mL) = 25] were harvested by centrifugation [5800 × g for 4 min at 25 °C] and 1 mL acetone was added to the cells and incubation with shaking for 30 min.…”

CyAbrB2 is a nucleoid-associated protein inSynechocystiscontrolling hydrogenase expression during fermentation

“…PFOR works coordinatedly with hydrogenase to reduce ferredoxin, which hydrogenase uses as an electron donor (7, 10). Despite their O 2 sensitivity, hydrogenase and PFOR work under mixotrophic (i.e., aerobic) conditions (11, 12). Therefore, uncontrolled expression of hox operon and nifJ may hamper metabolism under photosynthetic conditions or shorten the supply of inorganic cofactors.…”

“…Although hydrogenase and PFOR are O2 sensitive, they can work under aerobic conditions (10)(11)(12). Therefore, uncontrolled expression of hox operon and nifJ (coding gene of PFOR) may hamper metabolism under photosynthetic conditions.…”

Evidence for Electron Transfer from the Bidirectional Hydrogenase to the Photosynthetic Complex I (NDH-1) in the Cyanobacterium Synechocystis sp. PCC 6803

“…It has long been known that this type of hydrogenase is responsible for hydrogen evolution in the light (so-called photoH 2 ) that is produced when dark anaerobic cells are suddenly exposed to strong light [17]. Recently, we showed that the enzyme is also working as an electron valve when the cells are shifted to high light intensities under aerobic conditions [18]. It remains unclear how the enzyme performs this task but NADH formation from reduced ferredoxin and its subsequent oxidation in respiration are a possible route.…”