Nitrogenase activity and dark CO2 fixation in the lichen Peltigera aphthosa Willd

The lichen Peltigera aphthosa consists of a fungus and green alga (Coccomyxa) in the main thallus and of a Nostoc located in superficial packets, intermixed with fungus, called cephalodia. Dark nitrogenase activity (acetylene reduction) of lichen discs (of alga, fungus and Nostoc) and of excised cephalodia was sustained at higher rates and for longer than was the dark nitrogenase activity of the isolated Nostoc growing exponentially. Dark nitrogenase activity of the symbiotic Nostoc was supported by the catabolism of polyglucose accumulated in the ligh and which in darkness served to supply ATP and reductant. The decrease in glucose content of the cephalodia paralleled the decline in dark nitrogenase activity in the presence of CO2; in the absence of CO2 dark nitrogenase activity declined faster although the rate of glucose loss was similar in the presence and absence of CO2. Dark CO2 fixation, which after 30 min in darkness represented 17 and 20% of the light rates of discs and cephalodia, respectively, also facilitated dark nitrogenase activity. The isolated Nostoc, the Coccomyxa and the excised fungus all fixed CO2 in the dark; in the lichen most dark CO2 fixation was probably due to the fungus. Kinetic studies using discs or cephalodia showed highest initial incorporation of (14)CO2 in the dark in to oxaloacetate, aspartate, malate and fumarate; incorporation in to alanine and citrulline was low; incorporation in to sugar phosphates, phosphoglyceric acid and sugar alcohols was not significant. Substantial activities of the enzymes phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) and carbamoyl-phosphate synthase (EC 2.7.2.5 and 2.7.2.9) were detected but the activities of PEP carboxykinase (EC 4.1.1.49) and PEP carboxyphosphotransferase (EC 4.1.1.38) were negligible. In the dark nitrogenase activity by the cephalodia, but not by the free-living Nostoc, declined more rapidly in the absence than in the presence of CO2 in the gas phase. Exogenous NH 4 (+) inhibited nitrogenase activity by cephalodia in the dark especially in the absence of CO2 but had no effect in the light. The overall data suggest that in the lichen dark CO2 fixation by the fungus may provide carbon skeletons which accept NH 4 (+) released by the cyanobacterium and that in the absence of CO2, NH 4 (+) directly, or indirectly via a mechanism which involves glutamine synthetase, inhibits nitrogenase activity.