This title appears in the Scientific Report :
2003
Please use the identifier:
http://hdl.handle.net/2128/84 in citations.
Infrared spectroscopy of cytochrome c oxidase intermediate states
Infrared spectroscopy of cytochrome c oxidase intermediate states
Cytochrome c oxidase is a critical player in the process of cellular respiration, performing proton translocation across a lipid bilayer, coupled to the four-electron reduction of O$_{2}$ to H$_{2}$O. To accomplish this catalytic task, specific changes at the active site influence chemical and physi...
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Personal Name(s): | Nyquist, Rebecca M. (Corresponding author) |
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Contributing Institute: |
Biologische Strukturforschung; IBI-2 |
Imprint: |
Jülich
Forschungszentrum Jülich GmbH Zentralbibliothek, Verlag
2003
|
Physical Description: |
XIII, 178 p. |
Dissertation Note: |
Urbana-Champaign, Univ., Diss., 2002 |
Document Type: |
Book Dissertation / PhD Thesis |
Research Program: |
Neurowissenschaften |
Series Title: |
Berichte des Forschungszentrums Jülich
4009 |
Subject (ZB): | |
Link: |
OpenAccess |
Publikationsportal JuSER |
Cytochrome c oxidase is a critical player in the process of cellular respiration, performing proton translocation across a lipid bilayer, coupled to the four-electron reduction of O$_{2}$ to H$_{2}$O. To accomplish this catalytic task, specific changes at the active site influence chemical and physical changes throughout the protein, altering amino acid side-chain orientations, hydrogen bond lengths, and protonation states. Infrared spectroscopy is capable of monitoring these changes. In this thesis work, cytochrome c oxidase was specially prepared for perfusion-induced infrared difference spectroscopy. The resulting infrared difference spectra demonstrate that the side-chain of a key glutamic acid, E286 from $\textit{Rhodobacter sphaeroides}$, is protonated in both oxidized (O) and fully-reduced states with a pK$_{a}$ higher than 9.5. Also presented in this work are the first infrared difference spectra for O$_{2}$ bond-cleaved intermediate states $\textbf{P}$ and $\textbf{F}$. In addition, time-resolved infrared spectroscopy was used to study vibrational differences between intermediate states preceding O$_{2}$ binding, the one- and two-electron reduced states ($\textbf{E}$ and $\textbf{R}_{2}$, respectively). Taken together, the infrared difference spectra presented here demonstrate that the E286 side-chain is deprotonated in $\textbf{E}$ and $\textbf{P}$ but protonated in $\textbf{O}, \textbf{R}_{2}$, and $\textbf{F}$. This indicates that E286 transfers its proton in the $\textbf{O} \rightarrow \textbf{E}$ and $\textbf{R}_{2} \rightarrow \textbf{P}$ transitions; and that it accepts a proton in the $\textbf{E} \rightarrow \textbf{R}_{2}$ and $\textbf{P} \rightarrow \textbf{F}$ transitions. Also, a tyrosine residue, presumably the active site tyrosine Y288, was observed to be protonated in O and deprotonated in $\textbf{F}$. These results spark interpretation of mechanistic models as well as form the basis for future time-resolved infrared spectroscopic investigations. |