Protein control of the redox potential of the primary quinone acceptor in reactioncCenters from Rhodobacter sphaeroides.
The role of the protein environment in determining the redox midpoint potential (E(m)) of Q(A), the primary quinone of bacterial reaction centers, was investigated by mutation of isoleucine at position 265 of the M subunit in Rhodobacter sphaeroides. Isoleucine was changed to threonine, serine, and valine, yielding mutants M265IT, M265IS, and M265IV, respectively. All three mutants, with smaller residues replacing isoleucine, exhibited decreased binding affinities of the Q(A) site for various quinone analogues, consistent with an enlargement or loosening of the headgroup binding domain and a decrease in the van der Waals contact for small quinones. In all other respects, M265IV was like the wild type, but the polar mutants, M265IT and M265IS, had unexpectedly dramatic decreases in the redox midpoint potential of Q(A), resulting in faster rates of P(+)Q(A)(-) charge recombination. For both anthraquinone and native ubiquinone, the in situ E(m) of Q(A) was estimated to be approximately 100 and 85 mV lower in M265IT and M265IS, respectively. The effect on E(m)(Q(A)) indicates destabilization of the semiquinone or stabilization of the quinone. This is suggested to arise from either (i) electrostatic interaction between the partial charges or dipole of the residue hydroxyl group and the charge distribution of quinone and semiquinone states with particular influence near the C4 carbonyl group or (ii) from hydrogen bonding interactions between the hydroxyl oxygen and the N(delta)H of histidine M219, causing a weakening of the hydrogen bond to the C4 carbonyl. The rate of the first electron transfer (k(AB)(()(1)())) in the polar mutants was the same as in the wild type at low pH but decelerated at higher pH with altered pH dependence. The rate of the second electron transfer (k(AB)(()(2)())) was 3-4-fold greater than in the wild type over the whole pH range from 4 to 11, consistent with a larger driving force for electron transfer derived from the lower E(m) of Q(A).
Takahashi
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E
Department of Biochemistry and Center for Biophysics & Computational Biology, 419 Roger Adams Laboratory/MC-712, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, Illinois 61801, USA.
Wells
T A
TA
Wraight
C A
CA
D019943:Amino Acid Substitution / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D004579:Electron Transport / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007532:Isoleucine / Q000235:genetics; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D009285:Naphthoquinones / Q000737:chemistry; D010084:Oxidation-Reduction; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry* / Q000235:genetics; D011809:Quinones / Q000737:chemistry*; D012242:Rhodobacter sphaeroides / Q000737:chemistry* / Q000235:genetics; D012694:Serine / Q000235:genetics; D013912:Threonine / Q000235:genetics; D014451:Ubiquinone / Q000737:chemistry
D009285:Naphthoquinones; D045322:Photosynthetic Reaction Center Complex Proteins; D011809:Quinones; D007532:Isoleucine; D014451:Ubiquinone; D013912:Threonine; D012694:Serine