title: Role of the conserved arginine pair in proton and electron transfer in cytochrome C oxidase.
abstract: A hydrogen-bonded network is observed above the hemes in all of the high-resolution crystal structures of cytochrome oxidases. It includes water and a pair of arginines, R481 and R482 (Rhodobacter sphaeroides numbering), that interact directly with heme a and the heme a(3) propionates. The hydrogen-bonded network provides potential pathways for proton release. The arginines, and the backbone peptide bond between them, have also been proposed to form part of a facilitated electron transfer route between Cu(A) and heme a. Our studies show that mutations of R482 (K, Q, and A) and R481 (K) retain substantial activity and are able to pump protons, but at somewhat reduced rates and stoichiometries. A slowed rate of electron transfer from cytochrome c to Cu(A) suggests a change in the orientation of cytochrome c binding in all but the R to K mutants. The mutant R482P is more perturbed in its structure and is altered in the redox potential difference between heme a and Cu(A): +18 mV for R482P and +46 mV for the wild type (heme a - Cu(A)). The electron transfer rate between Cu(A) and heme a is also altered from 93000 s(-1) in the wild type to 50 s(-1) in the oxidized R482P mutant, reminiscent of changes observed in a Cu(A)-ligand mutant, H260N. In neither case is the approximately 2000-fold change in the rate accounted for by the altered redox potentials, suggesting that both cause a major modification in the path or reorganization energy of electron transfer.
authors.affiliation: Department of Biochemistry, Michigan State University, East Lansing, Michigan 48824-1319, USA.
authors.forename: Jie
authors.initials: J
authors.lastname: Qian
authors.forename: Denise A
authors.initials: DA
authors.lastname: Mills
authors.forename: Lois
authors.initials: L
authors.lastname: Geren
authors.forename: Keifei
authors.initials: K
authors.lastname: Wang
authors.forename: Curt W
authors.initials: CW
authors.lastname: Hoganson
authors.forename: Bryan
authors.initials: B
authors.lastname: Schmidt
authors.forename: Carrie
authors.initials: C
authors.lastname: Hiser
authors.forename: Gerald T
authors.initials: GT
authors.lastname: Babcock
authors.forename: Bill
authors.initials: B
authors.lastname: Durham
authors.forename: Francis
authors.initials: F
authors.lastname: Millett
authors.forename: Shelagh
authors.initials: S
authors.lastname: Ferguson-Miller
mesh_terms: D000409:Alanine / Q000235:genetics; D001120:Arginine / Q000235:genetics; D002384:Catalysis; D017124:Conserved Sequence / Q000235:genetics; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport / Q000235:genetics; D003576:Electron Transport Complex IV / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004789:Enzyme Activation / Q000235:genetics; D005973:Glutamine / Q000235:genetics; D006418:Heme / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008239:Lysine / Q000235:genetics; D008274:Magnesium / Q000737:chemistry; D016297:Mutagenesis, Site-Directed; D010782:Photolysis; D011392:Proline / Q000235:genetics; D017494:Proton Pumps / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012242:Rhodobacter sphaeroides / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D013056:Spectrophotometry, Ultraviolet
chemical_list: D017494:Proton Pumps; D005973:Glutamine; C027728:heme a; D006418:Heme; D001120:Arginine; D011392:Proline; D003576:Electron Transport Complex IV; D008274:Magnesium; D008239:Lysine; D000409:Alanine