title: The CO Photodissociation and Recombination Dynamics of the W172Y/F282T Ligand Channel Mutant of Rhodobacter sphaeroides aa3 Cytochrome c Oxidase.
abstract: In the ligand channel of the cytochrome c oxidase from Rhodobacter sphaeroides (Rs aa3 ) W172 and F282 have been proposed to generate a constriction that may slow ligand access to and from the active site. To explore this issue, the tryptophan and phenylalanine residues in Rs aa3 were mutated to the less bulky tyrosine and threonine residues, respectively, which occupy these sites in Thermus thermophilus (Tt) ba3 cytochrome oxidase. The CO photolysis and recombination dynamics of the reduced wild-type Rs aa3 and the W172Y/F282T mutant were investigated using time-resolved optical absorption spectroscopy. The spectral changes associated with the multiple processes are attributed to different conformers. The major CO recombination process (44 μs) in the W172Y/F282T mutant is ~500 times faster than the predominant CO recombination process in the wild-type enzyme (~23 ms). Classical dynamic simulations of the wild-type enzyme and double mutant showed significant structural changes at the active site in the mutant, including movement of the heme a3 ring-D propionate toward CuB and reduced binuclear center cavity volume. These structural changes effectively close the ligand exit pathway from the binuclear center, providing a basis for the faster CO recombination in the double mutant.
authors.affiliation: Department of Chemistry and Biochemistry, University of California, Santa Cruz, CA.
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Cassano
authors.affiliation: Center for Biophysics and Computational Biology, Department of Chemistry, University of Illinois at Urbana-Champaign, Urbana, IL.
authors.forename: Sylvia K
authors.initials: SK
authors.lastname: Choi
authors.affiliation: Department of Chemistry and Biochemistry, University of California, Santa Cruz, CA.
authors.forename: William
authors.initials: W
authors.lastname: McDonald
authors.affiliation: Department of Chemistry and Biochemistry, University of California, Santa Cruz, CA.
authors.forename: Istvan
authors.initials: I
authors.lastname: Szundi
authors.affiliation: Department of Chemistry and Biochemistry, University of California, Santa Cruz, CA.
authors.forename: Terra R
authors.initials: TR
authors.lastname: Villa Gawboy
authors.affiliation: Center for Biophysics and Computational Biology, Department of Chemistry, University of Illinois at Urbana-Champaign, Urbana, IL.
authors.forename: Robert B
authors.initials: RB
authors.lastname: Gennis
authors.affiliation: Department of Chemistry and Biochemistry, University of California, Santa Cruz, CA.
authors.forename: Ólöf
authors.initials: Ó
authors.lastname: Einarsdóttir
mesh_terms: D002248:Carbon Monoxide; D003198:Computer Simulation; D003576:Electron Transport Complex IV / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D007700:Kinetics; D008958:Models, Molecular; D009154:Mutation; D010782:Photolysis; D011487:Protein Conformation; D012242:Rhodobacter sphaeroides / Q000201:enzymology*
chemical_list: D002248:Carbon Monoxide; D003576:Electron Transport Complex IV