Kinetics of H+ ion binding by the P+Q(A)- state of bacterial photosynthetic reaction centers: Rate limitation within the protein

P. Maróti, Colin A. Wraight

Research output: Contribution to journalArticle

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Abstract

The kinetics of flash-induced H+ ion binding by isolated reaction centers (RCs) of Rhodobacter sphaeroides, strain R-26, were measured, using pH indicators and conductimetry, in the presence of terbutryn to block electron transfer between the primary and secondary quinones (Q(A) and Q(B)), and in the absence of exogenous electron donors to the oxidized primary donor, P+, i.e., the P+Q(A) state. Under these conditions, proton binding by RCs is to the protein rather than to any of the cofactors. After light activation to form P+Q(A), the kinetics of proton binding were monoexponential at all pH values studied. At neutral pH, the apparent bimolecular rate constant was close to the diffusional limit for proton transfer in aqueous solution (~1011 M-1 s-1), but increased significantly in the alkaline pH range (e.g., 2 x 1013 M-1 s-1 at pH 10). The average slope of the pH dependence was -0.4 instead of -1.0, as might be expected for a H+ diffusion-controlled process. High activation energy (0.54 eV at pH 8.0) and weak viscosity dependence showed that H+ ion uptake by RCs is not limited by diffusion. The salt dependence of the H+ ion binding rate and the pK values of the protonatable amino acid residues of the reaction center implicated surface charge influences, and Gouy-Chapman theory provided a workable description of the ionic effects as arising from modulation of the pH at the surface of the RC. Incubation in D20 caused small increases in the pKs of the protonatable groups and a small, pH (pD)- dependent slowing of the binding rate. The salt, pH, temperature, viscosity, and D2O dependences of the proton uptake by RCs in the P+Q(A) state were accounted for by three considerations: 1) parallel pathways of H+ delivery to the RC, contributing to the observed (net) H+ disappearance; 2) rate limitation of the protonation of target groups within the protein by conformational dynamics; and 3) electrostatic influences of charged groups in the protein, via the surface pH.

Original languageEnglish
Pages (from-to)367-381
Number of pages15
JournalBiophysical Journal
Volume73
Issue number1
Publication statusPublished - Jul 1997

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Photosynthetic Reaction Center Complex Proteins
Ions
Proteins
Protons
Viscosity
Salts
Electrons
Rhodobacter sphaeroides
Quinones
Static Electricity
Membrane Proteins

ASJC Scopus subject areas

  • Biophysics

Cite this

Kinetics of H+ ion binding by the P+Q(A)- state of bacterial photosynthetic reaction centers : Rate limitation within the protein. / Maróti, P.; Wraight, Colin A.

In: Biophysical Journal, Vol. 73, No. 1, 07.1997, p. 367-381.

Research output: Contribution to journalArticle

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abstract = "The kinetics of flash-induced H+ ion binding by isolated reaction centers (RCs) of Rhodobacter sphaeroides, strain R-26, were measured, using pH indicators and conductimetry, in the presence of terbutryn to block electron transfer between the primary and secondary quinones (Q(A) and Q(B)), and in the absence of exogenous electron donors to the oxidized primary donor, P+, i.e., the P+Q(A) state. Under these conditions, proton binding by RCs is to the protein rather than to any of the cofactors. After light activation to form P+Q(A), the kinetics of proton binding were monoexponential at all pH values studied. At neutral pH, the apparent bimolecular rate constant was close to the diffusional limit for proton transfer in aqueous solution (~1011 M-1 s-1), but increased significantly in the alkaline pH range (e.g., 2 x 1013 M-1 s-1 at pH 10). The average slope of the pH dependence was -0.4 instead of -1.0, as might be expected for a H+ diffusion-controlled process. High activation energy (0.54 eV at pH 8.0) and weak viscosity dependence showed that H+ ion uptake by RCs is not limited by diffusion. The salt dependence of the H+ ion binding rate and the pK values of the protonatable amino acid residues of the reaction center implicated surface charge influences, and Gouy-Chapman theory provided a workable description of the ionic effects as arising from modulation of the pH at the surface of the RC. Incubation in D20 caused small increases in the pKs of the protonatable groups and a small, pH (pD)- dependent slowing of the binding rate. The salt, pH, temperature, viscosity, and D2O dependences of the proton uptake by RCs in the P+Q(A) state were accounted for by three considerations: 1) parallel pathways of H+ delivery to the RC, contributing to the observed (net) H+ disappearance; 2) rate limitation of the protonation of target groups within the protein by conformational dynamics; and 3) electrostatic influences of charged groups in the protein, via the surface pH.",
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