Red-light effects sensitized by methylene blue on nitrate reductase from spinach (Spinacia Oleracea L.) leaves
1984
García-Mauriño Ruiz-Berdejo, Sofía | Echevarría Ruiz de Vargas, Cristina | Vargas Muñoz, María de los Ángeles | Aparicio, Pedro J. | Maldonado Ruiz, José María | Universidad de Sevilla. Departamento de Biología Vegetal y Ecología
Nitrate reductase from spinach (Spinacia oleracea L.) leaves, which had been inactivated in vitro by incubation with N A D H and cyanide, was fully reactivated in minutes when irradiated in anaerobic conditions with red light in the presence o f m ethylene blue. Both the rate and the extent of reactivation increased with light intensity ( 6 to 100 W - m “ 2) and dye concentration (1 to 10 jiM). On the contrary, photoreactivation was com pletely abolished when N A D H or ethylenediaminetetra-acetic acid were present during irradiation. W e propose that methylene blue, when photo excited, exhibits a redox potential positive enough to reoxidise the CN~-reduced molybdenum complex settled in the inactive enzyme, thus causing its reactivation. On the other hand, prolonged irradiation o f nitrate reductase, under air and in the presence o f methylene blue, promoted an oxygen-dependent irreversible inactivation o f the two partial activities of the enzyme. This inactivation was markedly enhanced in 77% deuterated water and greatly prevented by azide, which indicates that singlet oxygen is the species primarily involved in the photooxidative inactivation o f the enzyme
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