title: Functional characterization of the chloroplast ferric chelate oxidoreductase enzyme.
abstract: Iron (Fe) has an essential role in the biosynthesis of chlorophylls and redox cofactors, and thus chloroplast iron uptake is a process of special importance. The chloroplast ferric chelate oxidoreductase (cFRO) has a crucial role in this process but it is poorly characterized. To study the localization and mechanism of action of cFRO, sugar beet (Beta vulgaris cv Orbis) chloroplast envelope fractions were isolated by gradient ultracentrifugation, and their purity was tested by western blotting against different marker proteins. The ferric chelate reductase (FCR) activity of envelope fractions was studied in the presence of NAD(P)H (reductants) and FAD coenzymes. Reduction of Fe(III)-ethylenediaminetetraacetic acid was monitored spectrophotometrically by the Fe(II)-bathophenanthroline disulfonate complex formation. FCR activity, that is production of free Fe(II) for Fe uptake, showed biphasic saturation kinetics, and was clearly associated only to chloroplast inner envelope (cIE) vesicles. The reaction rate was > 2.5 times higher with NADPH than with NADH, which indicates the natural coenzyme preference of cFRO activity and its dependence on photosynthesis. FCR activity of cIE vesicles isolated from Fe-deficient plants also showed clear biphasic kinetics, where the KM of the low affinity component was elevated, and thus this component was down-regulated.
authors: Department of Plant Physiology and Molecular Plant Biology, Institute of Biology, Eötvös Loránd University, Pázmány P. sétány 1/C, Budapest, 1117, Hungary.
authors: Ádám
authors: Á
authors: Solti
authors: Department of Plant Physiology and Molecular Plant Biology, Institute of Biology, Eötvös Loránd University, Pázmány P. sétány 1/C, Budapest, 1117, Hungary.
authors: Brigitta
authors: B
authors: Müller
authors: Department of Plant Physiology and Molecular Plant Biology, Institute of Biology, Eötvös Loránd University, Pázmány P. sétány 1/C, Budapest, 1117, Hungary.
authors: Viktória
authors: V
authors: Czech
authors: Department of Plant Physiology and Molecular Plant Biology, Institute of Biology, Eötvös Loránd University, Pázmány P. sétány 1/C, Budapest, 1117, Hungary.
authors: Éva
authors: É
authors: Sárvári
authors: Department of Plant Physiology and Molecular Plant Biology, Institute of Biology, Eötvös Loránd University, Pázmány P. sétány 1/C, Budapest, 1117, Hungary.
authors: Ferenc
authors: F
authors: Fodor
mesh_terms: D027461:Beta vulgaris / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology; D002736:Chloroplasts / Q000187:drug effects / Q000201:enzymology*; D038181:FMN Reductase / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism; D007501:Iron / Q000494:pharmacology; D000090463:Iron Deficiencies; D010455:Peptides / Q000378:metabolism; D022161:Transport Vesicles / Q000187:drug effects / Q000378:metabolism
chemical_list: D010455:Peptides; D007501:Iron; D038181:FMN Reductase; C023903:ferric citrate iron reductase
keywords: chloroplast envelope membranes; ferric chelate oxidoreductase; ferric chelate reductase activity; iron deficiency; iron uptake