Introduction: Streptomyces coelicolor A3(2) is a filamentous actinobacterium that requires oxygen for growth and uses a branched respiratory chain to respire with O₂. A menaquinol:cytochrome bcc oxidoreductase and copper-aa₃-type cytochrome c oxidase supercomplex forms one branch, while the other includes a menaquinol:cytochrome bd oxidase. The formation of an electron-transferring supercomplex is necessary because S. coelicolor has a membrane-bound c-type cytochrome. Although S. coelicolor can only grow with oxygen, it also encodes three related respiratory nitrate reductases (Nar); however, these do not support growth but are presumed to help maintain a membrane potential when O₂ becomes limiting and nitrate is available. The Nar enzymes are synthesized and active in different stages of the complex developmental life cycle of the bacterium. Nar1 is exclusively present and active in spores, Nar2 mainly in exponentially growing mycelium and Nar3 in stationary mycelium. Nar1 activity depends on a functional cytochrome bcc-aa₃ supercomplex, but Nar2 activity in mycelium is not totally dependent on the respiratory supercomplex.

Objective: We wish to determine the mechanism underlying this differential dependence of Nar activity on the respiratory supercomplex at different developmental stages.

Methods: A combination of proteomic analyses, biochemical assays, use of defined nar mutants, physiological studies, including use of respiratory inhibitors, and determination of nitrate and oxygen consumption rates were used.

Results: Protein complex purification studies and proteomic analyses demonstrated that the respiratory supercomplexes differ between spores and mycelium. This finding was verified by determination of O₂ respiration rates combined with the use of respiratory inhibitors. The influence of nar1 and nar2 operon mutations on O₂ and nitrate reduction in spores and mycelium indicate direct electron transfer between Nar1 and the respiratory supercomplex in spores.

Conclusions: Our findings suggest that the cytochrome bcc- aa₃ supercomplex is differently organized in spores compared with mycelium. Nar1 appears to receive electrons directly from the cytochrome bcc-aa₃ supercomplex in spores, while Nar2 can functions as a menaquinol oxidase in mycelium. The ability of spores to switch rapidly between available electron acceptors (oxygen or nitrate) improves the chances of S. coelicolor spore survival.