Atp hydrolysis and electron transfer in the nitrogenase reaction with different combinations of the iron protein and the molybdenum-iron protein

Abstract

Purified iron protein (Fe protein) and molybdenum-iron protein (Mo-Fe protein) of nitrogenase from Clostridium pasteurianum were used for studies of ATP hydrolysis and electron transfer in the nitrogenase reaction. Total electron transfer was determined by measuring dithionite oxidation spectrophotometrically. Comparison with manometric determination of H 2 evolution showed that dithionite serves as a two electron donor. The studies of different combinations of the Fe protein and the Mo-Fe protein showed that excess Mo-Fe protein inhibited electron transfer but not ATP hydrolysis, whereas excess Fe protein did not inhibit either reaction. The ATP:2 electron ratios ranged from around 4 with excess Fe protein to more than 20 with excess Mo-Fe protein. With relatively low levels of Mo-Fe protein, reductant-independent ATP hydrolysis was around 6% of the reductant-dependent value, and with a large excess of Mo-Fe protein it increased to about 10%. We conclude that because the ATP:2 electron ratio varies with experimental conditions, no single value can be ascribed to this ratio.