title: Stigmatellin probes the electrostatic potential in the QB site of the photosynthetic reaction center.
abstract: The electrostatic potential in the secondary quinone (QB) binding site of the reaction center (RC) of the photosynthetic bacterium Rhodobacter sphaeroides determines the rate and free energy change (driving force) of electron transfer to QB. It is controlled by the ionization states of residues in a strongly interacting cluster around the QB site. Reduction of the QB induces change of the ionization states of residues and binding of protons from the bulk. Stigmatellin, an inhibitor of the mitochondrial and photosynthetic respiratory chain, has been proven to be a unique voltage probe of the QB binding pocket. It binds to the QB site with high affinity, and the pK value of its phenolic group monitors the local electrostatic potential with high sensitivity. Investigations with different types of detergent as a model system of isolated RC revealed that the pK of stigmatellin was controlled overwhelmingly by electrostatic and slightly by hydrophobic interactions. Measurements showed a high pK value (>11) of stigmatellin in the QB pocket of the dark-state wild-type RC, indicating substantial negative potential. When the local electrostatics of the QB site was modulated by a single mutation, L213Asp → Ala, or double mutations, L213Asp-L212Glu → Ala-Ala (AA), the pK of stigmatellin dropped to 7.5 and 7.4, respectively, which corresponds to a >210 mV increase in the electrostatic potential relative to the wild-type RC. This significant pK drop (ΔpK > 3.5) decreased dramatically to (ΔpK > 0.75) in the RC of the compensatory mutant (AA+M44Asn → AA+M44Asp). Our results indicate that the L213Asp is the most important actor in the control of the electrostatic potential in the QB site of the dark-state wild-type RC, in good accordance with conclusions of former studies using theoretical calculations or light-induced charge recombination assay.
authors.affiliation: Department of Biophysics, University of Szeged, Szeged, Hungary; Department of Physics and Astronomy, Vrije Universiteit Amsterdam, Amsterdam, The Netherlands. Electronic address: l.gerencser@vu.nl.
authors.forename: László
authors.initials: L
authors.lastname: Gerencsér
authors.affiliation: Department of Biophysics, University of Szeged, Szeged, Hungary.
authors.forename: Bogáta
authors.initials: B
authors.lastname: Boros
authors.affiliation: Laboratoire de Chimie Physique, University of Paris-Sud, Orsay, France.
authors.forename: Valerie
authors.initials: V
authors.lastname: Derrien
authors.affiliation: Biosciences Divisions, Argonne National Laboratory, Argonne, Illinois.
authors.forename: Deborah K
authors.initials: DK
authors.lastname: Hanson
authors.affiliation: Department of Biochemistry and Center for Biophysics and Computational Biology, University of Illinois, Urbana, Illinois.
authors.forename: Colin A
authors.initials: CA
authors.lastname: Wraight
authors.affiliation: Laboratoire de Chimie Physique, University of Paris-Sud, Orsay, France.
authors.forename: Pierre
authors.initials: P
authors.lastname: Sebban
authors.affiliation: Department of Biophysics, University of Szeged, Szeged, Hungary. Electronic address: pmaroti@sol.cc.u-szeged.hu.
authors.forename: Péter
authors.initials: P
authors.lastname: Maróti
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D016227:Benzoquinones / Q000378:metabolism; D001665:Binding Sites; D008969:Molecular Sequence Data; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011090:Polyenes / Q000737:chemistry / Q000494:pharmacology; D011485:Protein Binding; D012242:Rhodobacter sphaeroides / Q000201:enzymology; D055672:Static Electricity
chemical_list: D000900:Anti-Bacterial Agents; D016227:Benzoquinones; D045322:Photosynthetic Reaction Center Complex Proteins; D011090:Polyenes; C004532:quinone; C041573:stigmatellin