Anaerobically isolated endophytic vesicle clusters from root nodules of Alnusrubra Bong. were low in cellular energy charge (0.4 to 0.5), total adenylate pool (0.4 to 0.9 nmolùmg protein−1), and nitrogenase activity (0.1 to 0.3 nmolùmg protein−1). Upon addition of substrate (glucose, sucrose, isocitrate + isocitrate dehydrogenase + NADP), energy charge decreased by 2 to 20%, total adenylate pool increased by 3- to 11-fold, and nitrogenase activity increased by 50 to 500%. Only ATP and dithionite supported substantial nitrogenase activity and greatly elevated energy status. Treating vesicle clusters with macerase and cellulase to remove the host capsule facilitated substrate and oxygen uptake, resulting in a 240% increase in total adenylate pool, a 60% increase in energy charge, and a 730% increase in nitrogenase activity. Isolated vesicle clusters contained about 320 nmol cytochromes•g protein−1, with an average molar distribution of 17% cytochrome a-a3, 39% b, 22% c, and 21% o. No cytochrome P 450 was observed in vesicle clusters. The presence of cytochromes a-a3 and o may be a mechanism of oxygen tolerance whereby the nitrogenase in Frankia spp. can operate in an environment without the protection of leghaemoglobin.
