title: Isolation of novel PSII-LHCII megacomplexes from pea plants characterized by a combination of proteomics and electron microscopy.
abstract: In higher plants, photosystem II (PSII) is a multi-subunit pigment-protein complex embedded in the thylakoid membranes of chloroplasts, where it is present mostly in dimeric form within the grana. Its light-harvesting antenna system, LHCII, is composed of trimeric and monomeric complexes, which can associate in variable number with the dimeric PSII core complex in order to form different types of PSII-LHCII supercomplexes. Moreover, PSII-LHCII supercomplexes can laterally associate within the thylakoid membrane plane, thus forming higher molecular mass complexes, termed PSII-LHCII megacomplexes (Boekema et al. 1999a, in Biochemistry 38:2233-2239; Boekema et al. 1999b, in Eur J Biochem 266:444-452). In this study, pure PSII-LHCII megacomplexes were directly isolated from stacked pea thylakoid membranes by a rapid single-step solubilization, using the detergent n-dodecyl-α-D-maltoside, followed by sucrose gradient ultracentrifugation. The megacomplexes were subjected to biochemical and structural analyses. Transmission electron microscopy on negatively stained samples, followed by single-particle analyses, revealed a novel form of PSII-LHCII megacomplexes, as compared to previous studies (Boekema et al.1999a, in Biochemistry 38:2233-2239; Boekema et al. 1999b, in Eur J Biochem 266:444-452), consisting of two PSII-LHCII supercomplexes sitting side-by-side in the membrane plane, sandwiched together with a second copy. This second copy of the megacomplex is most likely derived from the opposite membrane of a granal stack. Two predominant forms of intact sandwiched megacomplexes were observed and termed, according to (Dekker and Boekema 2005 Biochim Biophys Acta 1706:12-39), as (C2S2)4and (C2S2+ C2S2M2)2megacomplexes. By applying a gel-based proteomic approach, the protein composition of the isolated megacomplexes was fully characterized. In summary, the new structural forms of isolated megacomplexes and the related modeling performed provide novel insights into how PSII-LHCII supercomplexes may bind to each other, not only in the membrane plane, but also between granal stacks within the chloroplast.
authors.affiliation: Applied Science and Technology Department - BioSolar Lab, Politecnico di Torino, Viale T. Michel 5, 15121, Alessandria, Italy.|Department of Biology, University of Padova, Via Ugo Bassi 58 B, 35121, Padova, Italy.
authors.forename: Pascal
authors.initials: P
authors.lastname: Albanese
authors.affiliation: School of Biological and Chemical Sciences, Queen Mary University of London, London, E1 4NS, UK.
authors.forename: Jon
authors.initials: J
authors.lastname: Nield
authors.affiliation: Center for Space Human Robotics IIT@POLITO, Istituto Italiano di Tecnologia, Corso Trento 21, 10129, Turin, Italy.
authors.forename: Jose Alejandro Muñoz
authors.initials: JA
authors.lastname: Tabares
authors.affiliation: Center for Space Human Robotics IIT@POLITO, Istituto Italiano di Tecnologia, Corso Trento 21, 10129, Turin, Italy.
authors.forename: Angelica
authors.initials: A
authors.lastname: Chiodoni
authors.affiliation: ISALIT-Department of Science and Technological Innovation, University of Eastern Piedmont, Viale T. Michel 11, 15121, Alessandria, Italy.|Department of Science and Technological Innovation, University of Eastern Piedmont, Viale T. Michel 11, 15121, Alessandria, Italy.
authors.forename: Marcello
authors.initials: M
authors.lastname: Manfredi
authors.affiliation: Department of Science and Technological Innovation, University of Eastern Piedmont, Viale T. Michel 11, 15121, Alessandria, Italy.
authors.forename: Fabio
authors.initials: F
authors.lastname: Gosetti
authors.affiliation: Department of Science and Technological Innovation, University of Eastern Piedmont, Viale T. Michel 11, 15121, Alessandria, Italy.
authors.forename: Emilio
authors.initials: E
authors.lastname: Marengo
authors.affiliation: Applied Science and Technology Department - BioSolar Lab, Politecnico di Torino, Viale T. Michel 5, 15121, Alessandria, Italy.
authors.forename: Guido
authors.initials: G
authors.lastname: Saracco
authors.affiliation: Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, London, SW7 2AZ, UK.
authors.forename: James
authors.initials: J
authors.lastname: Barber
authors.affiliation: Applied Science and Technology Department - BioSolar Lab, Politecnico di Torino, Viale T. Michel 5, 15121, Alessandria, Italy. cristina.pagliano@polito.it.
authors.forename: Cristina
authors.initials: C
authors.lastname: Pagliano
mesh_terms: D004591:Electrophoresis, Polyacrylamide Gel; D045342:Light-Harvesting Protein Complexes / Q000737:chemistry / Q000302:isolation & purification* / Q000502:physiology; D013058:Mass Spectrometry / Q000379:methods; D008854:Microscopy, Electron; D046529:Microscopy, Electron, Transmission; D018532:Pisum sativum / Q000737:chemistry / Q000502:physiology*; D045332:Photosystem II Protein Complex / Q000737:chemistry / Q000302:isolation & purification* / Q000502:physiology; D040901:Proteomics / Q000379:methods
chemical_list: D045342:Light-Harvesting Protein Complexes; D045332:Photosystem II Protein Complex
keywords: PSII-LHCII megacomplex; Photosystem II; Proteomics; Single-particle analysis; Transmission electron microscopy