Activity of Spore-Specific Respiratory Nitrate Reductase 1 of A3(2) Requires a Functional Cytochrome Oxidase Supercomplex.

Spores have strongly reduced metabolic activity and are produced during the complex developmental cycle of the actinobacterium Resting spores can remain viable for decades, yet little is known about how they conserve energy. It is known, however, that they can reduce either oxygen or nitrate using endogenous electron sources. uses either a cytochrome oxidase or a cytochrome oxidase supercomplex to reduce oxygen, while nitrate is reduced by Nar-type nitrate reductases, which typically oxidize quinol directly. Here, we show that in resting spores the Nar1 nitrate reductase requires a functional supercomplex to reduce nitrate. Mutants lacking the complete genetic locus encoding the supercomplex showed no Nar1-dependent nitrate reduction. Recovery of Nar1 activity was achieved by genetic complementation but only when the complete locus was reintroduced to the mutant strain. We could exclude that the dependence on the supercomplex for nitrate reduction was via regulation of nitrate transport. Moreover, the catalytic subunit, NarG1, of Nar1 was synthesized in the mutant, ruling out transcriptional control. Constitutive synthesis of Nar1 in mycelium revealed that the enzyme was poorly active in this compartment, suggesting that the Nar1 enzyme cannot act as a typical quinol oxidase. Notably, nitrate reduction by the Nar2 enzyme, which is active in growing mycelium, was not wholly dependent on the supercomplex for activity. Together, our data suggest that Nar1 functions together with the proton-translocating supercomplex to increase the efficiency of energy conservation in resting spores. forms spores that respire with either oxygen or nitrate, using only endogenous electron donors. This helps maintain a membrane potential and, thus, viability. Respiratory nitrate reductase (Nar) usually receives electrons directly from reduced quinone species; however, we show that nitrate respiration in spores requires a respiratory supercomplex comprising cytochrome oxidoreductase and oxidase. Our findings suggest that the Nar1 enzyme in the spore functions together with the proton-translocating supercomplex to help maintain the membrane potential more efficiently. Dissecting the mechanisms underlying this survival strategy is important for our general understanding of bacterial persistence during infection processes and of how bacteria might deal with nutrient limitation in the natural environment.