Hydrogen Bonds Involved in Binding the Qi-site Semiquinone in the bc1 Complex, Identified through Deuterium Exchange Using Pulsed EPR

Sergei A. Dikanov, Rimma I. Samoilova, Derrick R.J. Kolling, J. Todd Holland, Antony R. Crofts

Research output: Contribution to journalArticlepeer-review

Abstract

Exchangeable protons in the immediate neighborhood of the semiquinone (SQ) at the Qi-site of the bc1 complex (ubihydroquinone:cytochrome c oxidoreductase (EC 1.10.2.2)) from Rhodobacter sphaeroides have been characterized using electron spin echo envelope modulation (ESEEM) and hyperfine sublevel correlation spectroscopy (HYSCORE) and visualized by substitution of H2O by 2H2O. Three exchangeable protons interact with the electron spin of the SQ. They possess different isotropic and anisotropic hyperfine couplings that allow a clear distinction between them. The strength of interactions indicates that the protons are involved in hydrogen bonds with SQ. The hyperfine couplings differ from values typical for in-plane hydrogen bonds previously observed in model experiments. It is suggested that the two stronger couplings involve formation of hydrogen bonds with carbonyl oxygens, which have a significant out-of-plane character due to the combined influence of bulky substituents and the protein environment. These two hydrogen bonds are most probably to side chains suggested from crystallographic structures (His-217 and Asp-252 in R. sphaeroides). Assignment of the third hydrogen bond is more ambiguous but may involve either a bond between Asn-221 and a methoxy O-atom or a bond to water. The structural and catalytic roles of the exchangeable protons are discussed in the context of three high resolution crystallographic structures for mitochondrial bc1 complexes. Potential H-bonds, including those to water molecules, form a network connecting the quinone (ubiquinone) occupant and its ligands to the propionates of heme bH and the external aqueous phase. They provide pathways for exchange of protons within the site and with the exteriors, needed to accommodate the different hydrogen bonding requirements of different quinone species during catalysis.

Original languageEnglish (US)
Pages (from-to)15814-15823
Number of pages10
JournalJournal of Biological Chemistry
Volume279
Issue number16
DOIs
StatePublished - Apr 16 2004

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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