Proton uptake and pKa changes in the uncoupled Asn139Cys variant of cytochrome c oxidase

Ann Louise Johansson, Jens Carlsson, Martin Högbom, Jonathan P. Hosler, Robert B. Gennis, Peter Brzezinski

Research output: Contribution to journalArticle

Abstract

Cytochrome c oxidase (CytcO) is a membrane-bound enzyme that links electron transfer from cytochrome c to O2 to proton pumping across the membrane. Protons are transferred through specific pathways that connect the protein surface with the catalytic site as well as the proton input with the proton output sides. Results from earlier studies have shown that one site within the so-called D proton pathway, Asn139, located ∼10 Å from the protein surface, is particularly sensitive to mutations that uncouple the O 2 reduction reaction from the proton pumping activity. For example, none of the Asn139Asp (charged) or Asn139Thr (neutral) mutant CytcOs pump protons, although the proton-uptake rates are unaffected. Here, we have investigated the Asn139Cys and Asn139Cys/Asp132Asn mutant CytcOs. In contrast to other structural variants investigated to date, the Cys side chain may be either neutral or negatively charged in the experimentally accessible pH range. The data show that the Asn139Cys and Asn139Asp mutations result in the same changes of the kinetic and thermodynamic parameters associated with the proton transfer. The similarity is not due to introduction of charge at position 139, but rather introduction of a protonatable group that modulates the proton connectivity around this position. These results illuminate the mechanism by which CytcO couples electron transfer to proton pumping.

Original languageEnglish (US)
Pages (from-to)827-836
Number of pages10
JournalBiochemistry
Volume52
Issue number5
DOIs
StatePublished - Feb 5 2013

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ASJC Scopus subject areas

  • Biochemistry

Cite this

Johansson, A. L., Carlsson, J., Högbom, M., Hosler, J. P., Gennis, R. B., & Brzezinski, P. (2013). Proton uptake and pKa changes in the uncoupled Asn139Cys variant of cytochrome c oxidase. Biochemistry, 52(5), 827-836. https://doi.org/10.1021/bi301597a