The Effect of Activated Oxygen Species on Nitrogenase of Anabaena variabilis

The heterocyst-forming cyanobacterium Anabaena variabilis (ATCC 29413) is deficient in catalase activity, but especially heterocysts are rich in superoxide dismutase and peroxidases. It exhibits a light-dependent methylviologen-catalyzed oxygen uptake which is accompanied by a massive release o f hydrogen peroxide. Methylviologen caused a transient concentrationdependent inhibition o f nitrogenase ending in a complete loss o f activity. Suppression o f hy­ drogen peroxide accumulation by the photosynthesis inhibitor DCM U (diuron, 3-[3,4-dichlorophenyl]-l,l-dimethylurea), or by addition o f catalase prevented nitrogenase inhibition indi­ cating that methylviologen affects nitrogenase activity via hydrogen peroxide produced by photosynthetic electron transport. Immunospecific Western blotting o f cell extracts separated on SDS polyacrylamide gel electrophoresis showed that the transient inhibition o f nitrogenase is accompanied by rapid modification o f its Fe-protein followed by enzyme destruction. In a cell-free system the enzyme exhibited only moderate sensitivity towards hydrogen peroxide but was found susceptible against activated oxygen species generated by the xanthine/xanthine oxidase system and free iron. The results indicate that H-,0: itself has little direct effect on nitro­ genase, but endogenous peroxide produced by methylviologen in the light may initiate oxygen radical formation leading to nitrogenase modification and subsequently its destruction.