Flavin-mediated photoregulation of nitrate reductase: A key point of control in inorganic nitrogen photosynthetic metabolism

Abstract

The first step in the photosynthetic reduction of nitrate to ammonia, as carried out by plants and green algae, is catalyzed by the enzyme nitrate reductase. This enzyme serves an assimilatory function in which it catalyzes the exergonic two-electron reduction of nitrate to nitrite with electrons donated by pyridine nucleotides, which are in turn reduced at the end of the photosynthetic electron transport chain by electrons derived originally from water. Although the assimilation of nitrate by photosynthetic organisms appears to be regulated primarily at the nitrate uptake level, the nitrate reductase activity seems to be another key point of control for this pathway. In fact, nitrate reductase from eukaryotic organisms has been demonstrated to exist in two metabolically interconvertible forms, one oxidized/active and the other reduced/inactive. To account for such a redox interconversion of nitrate reductase, several mechanisms have been proposed. One of them is based on the characteristic photochemical properties of flavins, which can be reduced under illumination by a variety of electron donors, e.g. EDTA (or ethylenediaminetetraacetic acid), semicarbazide and amino acids, which, on the other hand, are unable to reduce flavins in the dark. This redox photoregulation of nitrate reductase activity has been studied in our laboratory using the enzyme from the green alga Monoraphidium braunii. Inactivation has been proposed to involve three steps: (i) reduction of flavins upon irradiation under reducing conditions, (ii) reduction of the enzyme-bound molybdenum centers, and (iii) formation of a stable complex with a nucleophilic agent such as cyanide, superoxide, or other. The reactivation process would also take place in three steps: (i) photoexcitation of flavins under oxidizing conditions, (ii) deoxidation of the regulatory centers by photoexcited flavins, and (iii) release of the nucleophilic agent bound to the enzyme. The redox conditions of the cells when flavins are photoexcited can thereby regulate nitrate reductase activity and hence nitrate assimilation.