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 O2reduction. These events trigge...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 2003-07, Vol.100 (15), p.8715-8720
Main Authors: Nyquist, Rebecca M., Heitbrink, Dirk, Bolwien, Carsten, Gennis, Robert B., Heberle, Joachim
Format: Article
Language:English
Subjects:
Active sites
Biochemistry
Biological Sciences
Biophysical Phenomena
Biophysics
Carbon Monoxide - chemistry
Catalysis
Cytochromes
Electron transfer
Electron Transport
Electron Transport Complex IV - chemistry
Electron Transport Complex IV - genetics
Electron Transport Complex IV - metabolism
Electrons
Enzymes
Glutamic Acid - chemistry
Infrared radiation
Models, Molecular
Mutagenesis, Site-Directed
Oxidases
Oxidation-Reduction
Protein Conformation
Protons
Rhodobacter sphaeroides - enzymology
Rhodobacter sphaeroides - genetics
Spectral bands
Spectrophotometry, Infrared
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Summary: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 O2reduction. 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,$R_2,\>P_m$, and F. These spectra reveal deprotonation of its key glutamic acid E286 in the E and in the Pmstates. 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.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.1530408100