title: In vitro analysis of the plastid terminal oxidase in photosynthetic electron transport.
abstract: The plastid terminal oxidase PTOX catalyzes the oxidation of plastoquinol (PQH2) coupled with the reduction of oxygen to water. In vivo PTOX is attached to the thylakoid membrane. PTOX is important for plastid development and carotenoid biosynthesis, and its role in photosynthesis is controversially discussed. To analyze PTOX activity in photosynthetic electron transport recombinant purified PTOX fused to the maltose-binding protein was added to photosystem II-enriched membrane fragments. These membrane fragments contain the plastoquinone (PQ) pool as verified by thermoluminescence. Experimental evidence for PTOX oxidizing PQH2 is demonstrated by following chlorophyll fluorescence induction. Addition of PTOX to photosystem II-enriched membrane fragments led to a slower rise, a lower level of the maximal fluorescence and an acceleration of the fluorescence decay. This effect was only observed at low light intensities indicating that PTOX cannot compete efficiently with the reduction of the PQ pool by photosystem II at higher light intensities. PTOX attached tightly to the membranes since it was only partly removable by membrane washings. Divalent cations enhanced the effect of PTOX on chlorophyll fluorescence compared to NaCl most likely because they increase connectivity between photosystem II centers and the size of the PQ pool. Using single turnover flashes, it was shown that the level of reactive oxygen species, generated by PTOX in a side reaction, increased when the spacing between subsequent double flashes was enlarged. This shows that PTOX generates reactive oxygen species under limited substrate availability.
authors.affiliation: Commissariat à l'Energie Atomique et aux Energies Alternatives (CEA) Saclay, Institut de Biologie et de Technologie de Saclay, Centre National de la Recherche Scientifique UMR 8221, Service de Bioénergétique, Biologie Structurale et Mécanisme, F-91191 Gif-sur-Yvette cedex, France.
authors.forename: Kathleen
authors.initials: K
authors.lastname: Feilke
authors.affiliation: Faculty of Biology, University of Freiburg, D-79104 Freiburg, Germany.
authors.forename: Qiuju
authors.initials: Q
authors.lastname: Yu
authors.affiliation: Faculty of Biology, University of Freiburg, D-79104 Freiburg, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Beyer
authors.affiliation: Commissariat à l'Energie Atomique et aux Energies Alternatives (CEA) Saclay, Institut de Biologie et de Technologie de Saclay, Centre National de la Recherche Scientifique UMR 8221, Service de Bioénergétique, Biologie Structurale et Mécanisme, F-91191 Gif-sur-Yvette cedex, France.
authors.forename: Pierre
authors.initials: P
authors.lastname: Sétif
authors.affiliation: Commissariat à l'Energie Atomique et aux Energies Alternatives (CEA) Saclay, Institut de Biologie et de Technologie de Saclay, Centre National de la Recherche Scientifique UMR 8221, Service de Bioénergétique, Biologie Structurale et Mécanisme, F-91191 Gif-sur-Yvette cedex, France. Electronic address: anja.krieger-liszkay@cea.fr.
authors.forename: Anja
authors.initials: A
authors.lastname: Krieger-Liszkay
mesh_terms: D002734:Chlorophyll / Q000378:metabolism; D004579:Electron Transport; D005453:Fluorescence; D066298:In Vitro Techniques; D010088:Oxidoreductases / Q000378:metabolism*; D010788:Photosynthesis; D018087:Plastids
chemical_list: D002734:Chlorophyll; D010088:Oxidoreductases
keywords: Chlorophyll fluorescence; Photosynthetic electron transport; Plastid terminal oxidase; Reactive oxygen species