Nitrite reduction in barley-root plastids: Dependence on NADPH coupled with glucose-6-phosphate and 6-phosphogluconate dehydrogenases, and possible involvement of an electron carrier and a diaphorase

Abstract

Plastids from roots of barley (Hordeum vulgare L.) seedlings were isolated by discontinuous Percoll-gradient centrifugation. Coinciding with the peak of nitrite reductase (NiR; EC 1.7.7.1, a marker enzyme for plastids) in the gradients was a peak of a glucose-6-phosphate (Glc6P) and NADP(+)-linked nitrite-reductase system. High activities of phosphohexose isomerase (EC 5.3.1.9) and phosphoglucomutase (EC 2.7.5.1) as well as glucose-6-phosphate dehydrogenase (Glc6PDH; EC 1.1.1.49) and 6-phosphogluconate dehydrogenase (6PGDH; EC 1.1.1.44) were also present in the isolated plastids. Thus, the plastids contained an overall electron-transport system from NADPH coupled with Glc6PDH and 6PGDH to nitrite, from which ammonium is formed stoichiometrically. However, NADPH alone did not serve as an electron donor for nitrite reduction, although NADPH with Glc6P added was effective. Benzyl and methyl viologens were enzymatically reduced by plastid extract in the presence of Glc6P+ NADP(+). When the plastids were incubated with dithionite, nitrite reduction took place, and ammonium was formed stoichiometrically. The results indicate that both an electron carrier and a diaphorase having ferredoxin-NADP(+) reductase activity are involved in the electron-transport system of root plastids from NADPH, coupled with Glc6PDH and 6PGDH, to nitrite.

NADPH Thioredoxin Reductase C Is Localized in Plastids of Photosynthetic and Nonphotosynthetic Tissues and Is Involved in Lateral Root Formation in Arabidopsis

“…Thus, NTRC may constitute a direct pathway for redox homeostasis in heterotrophic plastids (Figure 10). However, transfer of electrons from NADPH to Fd, catalyzed by FNR, is also possible in heterotrophic plastids, as suggested by the high expression of RFNR2 in roots ( Figure 9A), in agreement with the previous report of a root-specific form of this enzyme (Oji et al, 1985). Though at a lower level than in leaves, qPCR analyses showed the expression in roots of genes encoding Fd, the catalytic and regulatory subunits of FTR and TRX f1 and TRX f2 (Figures 9B to 9D), a pattern confirmed by the Genevestigator data (see Supplemental Table 2 online).…”

Structural and functional characterization of ferredoxin‐NADP+‐oxidoreductase using knock‐out mutants of Arabidopsis

“…In higher plants, a small multigene family encodes two distinct forms of FNR (Morigasaki et al, 1993). The root form of FNR functions in non-photosynthetic plastids mediating electrons from NADPH, originating from the cytosolic glucose 6-phosphate via the oxidative pentose phosphate pathway, to the root-specific, nitrate-induced form of ferredoxin (Oji et al, 1985). Reduced ferredoxin, in turn, donates electrons for nitrite reduction by nitrite reductase, or for many other ferredoxin-dependent enzymes involved in assimilation of, for example, nitrogen and sulfur (Neuhaus and Emes, 2000).…”

Ferredoxin and Ferredoxin-NADP Reductase from Photosynthetic and Nonphotosynthetic Tissues of Tomato

“…15), these proteins were purified and characterized from roots ofboth radish (14,25) and spinach (15) and from etiolated bean sprouts (6,7). In these tissues, ferredoxin and FNR probably support ferredoxin-dependent biosynthetic processes, such as nitrogen assimilation, that take place in amyloplasts and proplastids (17,24 (12). FNR was assayed by the same method except that FNR was omitted from the reaction mixture and was replaced with excess spinach leaf ferredoxin.…”