title: Identification of pH-sensing Sites in the Light Harvesting Complex Stress-related 3 Protein Essential for Triggering Non-photochemical Quenching in Chlamydomonas reinhardtii.
abstract: Light harvesting complex stress-related 3 (LHCSR3) is the protein essential for photoprotective excess energy dissipation (non-photochemical quenching, NPQ) in the model green algaChlamydomonas reinhardtii Activation of NPQ requires low pH in the thylakoid lumen, which is induced in excess light conditions and sensed by lumen-exposed acidic residues. In this work we have used site-specific mutagenesisin vivoandin vitrofor identification of the residues in LHCSR3 that are responsible for sensing lumen pH. Lumen-exposed protonatable residues, aspartate and glutamate, were mutated to asparagine and glutamine, respectively. By expression in a mutant lacking all LHCSR isoforms, residues Asp(117), Glu(221), and Glu(224)were shown to be essential for LHCSR3-dependent NPQ induction inC. reinhardtii Analysis of recombinant proteins carrying the same mutations refoldedin vitrowith pigments showed that the capacity of responding to low pH by decreasing the fluorescence lifetime, present in the wild-type protein, was lost. Consistent with a role in pH sensing, the mutations led to a substantial reduction in binding the NPQ inhibitor dicyclohexylcarbodiimide.
authors.affiliation: From the Department of Biotechnology, University of Verona, Strada Le Grazie, I-37134 Verona, Italy.
authors.forename: Matteo
authors.initials: M
authors.lastname: Ballottari
authors.affiliation: the Howard Hughes Medical Institute, Department of Plant and Microbial Biology, University of California, Berkeley, California 94720-3102.
authors.forename: Thuy B
authors.initials: TB
authors.lastname: Truong
authors.affiliation: the Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, and the Graduate Group in Applied Science and Technology, University of California, Berkeley, California 94720.
authors.forename: Eleonora
authors.initials: E
authors.lastname: De Re
authors.affiliation: the Howard Hughes Medical Institute, Department of Plant and Microbial Biology, University of California, Berkeley, California 94720-3102, the Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, and.
authors.forename: Erika
authors.initials: E
authors.lastname: Erickson
authors.affiliation: From the Department of Biotechnology, University of Verona, Strada Le Grazie, I-37134 Verona, Italy, the Sorbonne Universités, UPMC Univ-Paris 6, CNRS, UMR 7238, Laboratoire de Biologie Computationnelle et Quantitative, 15 rue de l'Ecole de Médecine, 75006 Paris, France.
authors.forename: Giulio R
authors.initials: GR
authors.lastname: Stella
authors.affiliation: the Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, and the Graduate Group in Applied Science and Technology, University of California, Berkeley, California 94720 the Department of Chemistry, Hildebrand B77, University of California, Berkeley, California 94720-1460.
authors.forename: Graham R
authors.initials: GR
authors.lastname: Fleming
authors.affiliation: From the Department of Biotechnology, University of Verona, Strada Le Grazie, I-37134 Verona, Italy, roberto.bassi@univr.it.
authors.forename: Roberto
authors.initials: R
authors.lastname: Bassi
authors.affiliation: the Howard Hughes Medical Institute, Department of Plant and Microbial Biology, University of California, Berkeley, California 94720-3102, the Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, and niyogi@berkeley.edu.
authors.forename: Krishna K
authors.initials: KK
authors.lastname: Niyogi
mesh_terms: D016825:Chlamydomonas reinhardtii / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D045342:Light-Harvesting Protein Complexes / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense; D020524:Thylakoids / Q000235:genetics / Q000378:metabolism*
chemical_list: D045342:Light-Harvesting Protein Complexes
keywords: fluorescence; non-photochemical quenching; photoprotection; photosynthesis; photosynthetic pigment; photosystem II; plant biochemistry
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