47] Nitrogen and hydrogen metabolism: Induction and measurement

Abstract

This chapter summarizes some comparatively simple routine assays for the measurement of both nitrogen fixation and hydrogen gas exchange in intact cyanobacteria. Special attention is given to the phenomena involved in the induction of nitrogenase activity in heterocystous and nonheterocystous filamentous cyanobacteria. Many nonheterocystous cyanobacteria are also able to fix dinitrogen, when incubated under microaerobic conditions. Nitrogenase activity is conveniently measured using the acetylene reduction assay. This method requires a gas chromatograph (GC) equipped with a Porapak column, a flame-ionizing detector and acetylene (C 2 H 2 ) as substrate. Acetylene, however, may often not be available in pure form. Contamination by methane is not critical for the nitrogenase reaction, but the methane is detected in the assay because it elutes from the GC column with a retention time shorter than that for ethylene. Hydrogen gas exchange is closely related to nitrogenase activity, because hydrogen formation is an obligatory side reaction of nitrogen fixation. Hydrogen produced by nitrogenase is generally taken up again by a hydrogenase, so that net hydrogen evolution by nitrogen-fixing cyanobacteria is hardly observed (at least under aerobic conditions).