title: Hydrogen bonding and spin density distribution in the Qb semiquinone of bacterial reaction centers and comparison with the Qa site.
abstract: In the photosynthetic reaction center from Rhodobacter sphaeroides, the primary (Q(A)) and secondary (Q(B)) electron acceptors are both ubiquinone-10, but with very different properties and functions. To investigate the protein environment that imparts these functional differences, we have applied X-band HYSCORE, a 2D pulsed EPR technique, to characterize the exchangeable protons around the semiquinone (SQ) in the Q(A) and Q(B) sites, using samples of (15)N-labeled reaction centers, with the native high spin Fe(2+) exchanged for diamagnetic Zn(2+), prepared in (1)H(2)O and (2)H(2)O solvent. The powder HYSCORE method is first validated against the orientation-selected Q-band ENDOR study of the Q(A) SQ by Flores et al. (Biophys. J.2007, 92, 671-682), with good agreement for two exchangeable protons with anisotropic hyperfine tensor components, T, both in the range 4.6-5.4 MHz. HYSCORE was then applied to the Q(B) SQ where we found proton lines corresponding to T ≈ 5.2, 3.7 MHz and T ≈ 1.9 MHz. Density functional-based quantum mechanics/molecular mechanics (QM/MM) calculations, employing a model of the Q(B) site, were used to assign the observed couplings to specific hydrogen bonding interactions with the Q(B) SQ. These calculations allow us to assign the T = 5.2 MHz proton to the His-L190 N(δ)H···O(4) (carbonyl) hydrogen bonding interaction. The T = 3.7 MHz spectral feature most likely results from hydrogen bonding interactions of O1 (carbonyl) with both Gly-L225 peptide NH and Ser-L223 hydroxyl OH, which possess calculated couplings very close to this value. The smaller 1.9 MHz coupling is assigned to a weakly bound peptide NH proton of Ile-L224. The calculations performed with this structural model of the Q(B) site show less asymmetric distribution of unpaired spin density over the SQ than seen for the Q(A) site, consistent with available experimental data for (13)C and (17)O carbonyl hyperfine couplings. The implications of these interactions for Q(B) function and comparisons with the Q(A) site are discussed.
authors.affiliation: Center for Biophysics & Computational Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
authors.forename: Erik
authors.initials: E
authors.lastname: Martin
authors.forename: Rimma I
authors.initials: RI
authors.lastname: Samoilova
authors.forename: Kupala V
authors.initials: KV
authors.lastname: Narasimhulu
authors.forename: Tzu-Jen
authors.initials: TJ
authors.lastname: Lin
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: O'Malley
authors.forename: Colin A
authors.initials: CA
authors.lastname: Wraight
authors.forename: Sergei A
authors.initials: SA
authors.lastname: Dikanov
mesh_terms: D016227:Benzoquinones / Q000737:chemistry* / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D006860:Hydrogen Bonding; D008958:Models, Molecular; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D011522:Protons; D011789:Quantum Theory; D012242:Rhodobacter sphaeroides
chemical_list: D016227:Benzoquinones; D045322:Photosynthetic Reaction Center Complex Proteins; D011522:Protons; C025232:semiquinone radicals