title: Biochemical characterization and electron-transfer reactions of sym1, a Rhodobacter capsulatus reaction center symmetry mutant which affects the initial electron donor.
abstract: A 51 bp section of the Rhodobacter capsulatus photosynthetic reaction center M subunit gene (nucleotides M562-M612 of the pufM structural sequence) encoding amino acids M187-M203 was replaced by the homologous region of the L subunit gene. This resulted in the symmetrization of much of the amino acid environment of the reaction center initial electron donor, P. This is the first in a series of large-scale symmetry mutations and is referred to as sym1. The sym1 mutant was able to grow photosynthetically, indicating that reaction center function was largely intact. Isolated reaction centers showed an approximately 10-nm blue shift in the QY band of P. The standard free energy change between P* and P+BphA- determined from analysis of the long-lived fluorescence from quinone-reduced reaction centers decreased from about -120 meV in the wild-type to about -75 meV in the sym1 mutant. A 65-70% quantum yield of electron transfer from P* to P+QA- was observed, most of the yield loss occurring between P* and P+BphA-. The decay of the stimulated emission from P* was about 3-fold slower in this mutant than in the wild-type. Time-resolved spectral analysis of the charge-separated intermediates formed in sym1 reaction centers indicated that the major product was P+BphA-. A model-dependent analysis of the observed rates and electron-transfer yields gave the following microscopic rate constants for sym1 reaction centers (wild-type values under the same conditions are given in parentheses): [formula: see text] Analysis of the sym1 mutant, mutants near P made by other groups, and interspecies variation of amino acids in the vicinity of P suggests that the protein asymmetry in the environment of the initial electron donor is important for optimizing the rate and yield of electron transfer, but is not strictly required for overall reaction center function.
authors.affiliation: Department of Chemistry and Biochemistry, Arizona State University, Tempe 85287-1604.
authors.forename: A K
authors.initials: AK
authors.lastname: Taguchi
authors.forename: J W
authors.initials: JW
authors.lastname: Stocker
authors.forename: R G
authors.initials: RG
authors.lastname: Alden
authors.forename: T P
authors.initials: TP
authors.lastname: Causgrove
authors.forename: J M
authors.initials: JM
authors.lastname: Peloquin
authors.forename: S G
authors.initials: SG
authors.lastname: Boxer
authors.forename: N W
authors.initials: NW
authors.lastname: Woodbury
mesh_terms: D000595:Amino Acid Sequence; D020130:Bacterial Chromatophores / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D004579:Electron Transport; D005798:Genes, Bacterial; D007700:Kinetics; D046911:Macromolecular Substances; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D045322:Photosynthetic Reaction Center Complex Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D010957:Plasmids; D011487:Protein Conformation; D011789:Quantum Theory; D016354:Rhodobacter capsulatus / Q000235:genetics* / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013053:Spectrophotometry
chemical_list: D046911:Macromolecular Substances; D045322:Photosynthetic Reaction Center Complex Proteins