title: Early folding events during light harvesting complex II assembly in vitro monitored by pulsed electron paramagnetic resonance.
abstract: Efficient energy transfer in the major light harvesting complex II (LHCII) of green plants is facilitated by the precise alignment of pigments due to the protein matrix they are bound to. Much is known about the import of the LHCII apoprotein into the chloroplast via the TOC/TIC system and its targeting to the thylakoid membrane but information is sparse about when and where the pigments are bound and how this is coordinated with protein folding. In vitro, the LHCII apoprotein spontaneously folds and binds its pigments if the detergent-solubilized protein is combined with a mixture of chlorophylls a and b and carotenoids. In the present work, we employed this approach to study apoprotein folding and pigment binding in a time-resolved manner by using pulsed electron paramagnetic resonance (EPR). Intra-molecular distances were measured before folding, after 255 ms and 40 s folding time in the absence of cryoprotectant, and in the fully folded and assembled LHCII. In accordance with earlier results, the most of the folding of the three membrane-spanning alpha helices precedes their apposition into the final tertiary structure. However, their formation follows different kinetics, partially extending into the final phase of LHCII formation during which much of the condensation of the pigment-protein structure occurs, presumably governed by the binding of chlorophyll b. A rough timetable is proposed to sort partial events into the LHCII formation process.
authors.affiliation: Department of General Botany, Johannes Gutenberg-University, 55128 Mainz, Germany.
authors.forename: Niklas
authors.initials: N
authors.lastname: Fehr
authors.affiliation: Department of Physical Chemistry, ETH Zurich, Vladimir Prelog-Weg 2, CH-8093, Zürich, Switzerland.
authors.forename: Inés
authors.initials: I
authors.lastname: García-Rubio
authors.affiliation: Department of Physical Chemistry, ETH Zurich, Vladimir Prelog-Weg 2, CH-8093, Zürich, Switzerland.
authors.forename: Gunnar
authors.initials: G
authors.lastname: Jeschke
authors.affiliation: Department of General Botany, Johannes Gutenberg-University, 55128 Mainz, Germany. Electronic address: paulsen@uni-mainz.de.
authors.forename: Harald
authors.initials: H
authors.lastname: Paulsen
mesh_terms: D001059:Apoproteins / Q000737:chemistry / Q000378:metabolism; D002734:Chlorophyll / Q000737:chemistry / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D004735:Energy Transfer; D007700:Kinetics; D045342:Light-Harvesting Protein Complexes / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D018532:Pisum sativum / Q000378:metabolism; D045332:Photosystem II Protein Complex / Q000737:chemistry* / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary
chemical_list: D001059:Apoproteins; D045342:Light-Harvesting Protein Complexes; D045332:Photosystem II Protein Complex; D010940:Plant Proteins; D002734:Chlorophyll
keywords: EPR; LHCII; Membrane protein; Time-resolved folding