Direct observation of protonation reactions during the catalytic cycle of cytochrome c oxidase.
Cytochrome c oxidase, the terminal protein in the respiratory chain, converts oxygen into water and helps generate the electrochemical gradient used in the synthesis of ATP. The catalytic action of cytochrome c oxidase involves electron transfer, proton transfer, and O2 reduction. These events trigger specific molecular changes at the active site, which, in turn, influence changes throughout the protein, including alterations of amino acid side chain orientations, hydrogen bond patterns, and protonation states. We have used IR difference spectroscopy to investigate such modulations for the functional intermediate states E, R2,Pm, and F. These spectra reveal deprotonation of its key glutamic acid E286 in the E and in the Pm states. The consecutive deprotonation and reprotonation of E286 twice within one catalytic turnover illustrates the role of this residue as a proton shuttle. In addition, the spectra point toward deprotonation of a redox-active tyrosine, plausibly Y288, in the F intermediate. Structural insights into the molecular mechanism of catalysis based on the subtle molecular changes observed with IR difference spectroscopy are discussed.
Nyquist
Rebecca M
RM
Forschungszentrum Jülich, IBI-2: Structural Biology, 52425 Jülich, Germany.
Heitbrink
Dirk
D
Bolwien
Carsten
C
Gennis
Robert B
RB
Heberle
Joachim
J
D055592:Biophysical Phenomena; D001703:Biophysics; D002248:Carbon Monoxide / Q000737:chemistry; D002384:Catalysis; D004579:Electron Transport; D003576:Electron Transport Complex IV / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018698:Glutamic Acid / Q000737:chemistry; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010084:Oxidation-Reduction; D011487:Protein Conformation; D011522:Protons; D012242:Rhodobacter sphaeroides / Q000201:enzymology* / Q000235:genetics; D013055:Spectrophotometry, Infrared
D011522:Protons; D018698:Glutamic Acid; D002248:Carbon Monoxide; D003576:Electron Transport Complex IV