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  • Glyceraldehyde 3-phosphate dehydrogenase  (1)
  • antimycin A  (1)
  • 1
    ISSN: 1432-2048
    Keywords: Key words:Capsicum (fruit plastids) ; Dark metabolism ; Glyceraldehyde 3-phosphate dehydrogenase ; Malate dehydrogenase ; Malate valve ; Spinacia (chloroplasts)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract. Chloroplasts isolated from spinach (Spinacia oleracea L.) leaves and green sweet-pepper (Capsicum annuum L. var. grossum (L.) Sendt.) fruits contain NADP-dependent malate dehydrogenase (MDH; EC 1.1.1.82) and the bispecific NAD(P)-glyceraldehyde 3-phosphate dehydrogenase (GAPDH; EC 1.2.1.13). The NADP-dependent MDH and GAPDH are activated in the light, and inactive in the dark. We found that chloroplasts possess additional NAD-dependent MDH activity which is, like the NAD-dependent GAPDH activity, not influenced by light. In heterotrophic chromoplasts from red sweet-pepper fruits, the NADP-dependent MDH and the NAD(P)-GAPDH isoenzymes disappear during the developmental transition and only NAD-specific isoforms are found. Spinach chloroplasts contain both NAD/H and NADP/H at significant concentrations. Measurements of the pyridine dinucleotide redox states, performed under dark and various light conditions, indicate that NAD(H) is not involved in electron flow in the light. To analyze the contribution of NAD(H)-dependent reactions during dark metabolism, plastids from spinach leaves or green and red sweet-pepper fruits were incubated with dihydroxyacetone phosphate (DHAP). Exogenously added DHAP was oxidized into 3-phosphoglycerate by all types of plastids only in the presence of oxaloacetate, but not with nitrite or in the absence of added electron acceptors. We conclude that the NAD-dependent activity of GAPDH is essential in the dark to produce the ATP required for starch metabolism; excess electrons produced during triose-phosphate oxidation can selectively be used by NAD-MDH to form malate. Thus NADPH produced independently in the oxidative pentose-phosphate pathway will remain available for reductive processes inside the plastids.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1573-5079
    Keywords: antimycin A ; chloroplasts ; hydrogen peroxide ; malate valve ; redox state
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Electron fluxes in isolated intact spinach chloroplasts were analyzed under saturating light and under optimal CO2 and Pi supply. When CO2 assimilation was the only ATP- and NADPH-consuming reaction, the ΔpH decreased and the chloroplasts showed clear evidence of over-reduction. This suggested that additional electron flow is required in order to maintain the ΔpH and the stromal NADPH/ATP ratio. The additional electron flow may be cyclic electron transport around Photosystem I and linear electron transport towards either oxaloacetate or O2. The contributions of, and the interrelationships between, these three electron transfer pathways were analyzed by following the reactions of chloroplasts in their presence or absence, and by monitoring to what extent they were able to compensate for each other. Inhibition of cyclic electron flow by antimycin A caused strong over-reduction and decreased the ΔpH. Only oxaloacetate, but not O2, was able to restore photosynthesis. In the presence of H2O2, there was a rapid build-up of a high ΔpH, and the reduction of any other electron acceptor was prevented. It is concluded that the different electron acceptors in the stroma are organized in a hierarchical manner; this allows electron flux towards CO2 and nitrite reduction to proceed without any competition for electrons, and any excess electrons to be taken by these additional non-assimilatory pathways. Hence, the ΔpH is maintained at the required level and over-reduction of the electron transport chain and the stromal redox components is avoided.
    Type of Medium: Electronic Resource
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