It has been shown that some aerobic, unicellular, diazotrophic cyanobacteria temporally separate photosynthetic 02 evolution and oxygen-sensitive N2 fixation. Cyanothece sp. ATCC strain 51142 is an aerobic, unicellular, diazotrophic cyanobacterium that fixes N2 during discrete periods of its cell cycle. When the bacteria are maintained under diurnal light-dark cycles, N2 fixation occurs in the darlk Similar cycling is observed in continuous light, implicating a circadian rhythm. Under N2-fixing conditions, large inclusion granules form between the thylakoid membranes. Maximum granulation, as observed by electron microscopy, occurs before the onset of N2 fixation, and the granules decrease in number during the period of N2 fixation.The granules can be purified from cell homogenates by differential centrifugation. Biochemical analyses of the granules indicate that these structures are primarily carbohydrate, with some protein. Further analyses of the carbohydrate have shown that it is a glucose polymer with some characteristics of glycogen. It is proposed that N2 fixation is driven by energy and reducing power stored in these inclusion granules. Cyanothece sp. strain ATCC 51142 represents an excellent experimental organism for the study of the protective mechanisms of nitrogenase, metabolic events in cyanobacteria under normal and stress conditions, the partitioning of resources between growth and storage, and biological rhythms.Some cyanobacteria use energy acquired through oxygenic photosynthesis to drive N2 fixation. Nitrogenase, the enzyme that catalyzes the six-electron reduction of dinitrogen to ammonia, is notoriously sensitive to molecular oxygen, so much so that it seems incompatible with the oxygen-evolving nature of photosynthetic organisms. However, cyanobacteria have devised remarkably elegant methods of protecting nitrogenase from contacting oxygen, including spatial separation, temporal separation, and efficient enzyme systems that destroy reactive 02 by-products (8).Aerobic, unicellular, diazotrophic cyanobacteria exhibit temporal separation of oxygenic photosynthetic activities and oxygen-sensitive N2 fixation (8) Dinitrogen fixation is an energy-demanding process, requiring several ATPs per molecule of combined nitrogen produced (41). Thus, for N2 fixation to continue in the dark, without light-driven energy production or exogenous nutrient sources, the cells must utilize internal stores of energy and reducing power. In fact, it has been shown that, under diazotrophic conditions, the aerobic, unicellular cyanobacteria accumulate large quantities of carbohydrates (9, 24-26). These pools of photosynthate accumulate during a phase of rapid CO2 fixation in the light and are then apparently utilized during the dark phase to drive N2 fixation (and presumably for maintenance energy). Work with both Synechococcus sp. strain Miami BG 043511 (24, 25) and Gloeothece spp. (9,26) has demonstrated that carbohydrates accumulate toward the end of the light phase and dissipate in the dark in cultures grown diazot...