Photosynthetic water oxidation by photosystem II is mediated by a Mn4 cluster, a cofactor X still chemically ill-defined, and a tyrosine, YZ (D1-Tyr161). Before the final reaction with water proceeds to yield O2 (transition S4-->S0), two oxidizing equivalents are stored on Mn4 (S0-->S1-->S2), a third on X (S2-->S3), and a forth on YZ(S3-->S4). It has been proposed that YZ functions as a pure electron transmitter between Mn4X and P680, or, more recently, that it acts as an abstractor of hydrogen from bound water. We scrutinized the coupling of electron and proton transfer during the oxidation of YZ in PSII core particles with intact or impaired oxygen-evolving capacity. The rates of electron transfer to P680+, of electrochromism, and of pH transients were determined as a function of the pH, the temperature, and the H/D ratio. In oxygen-evolving material, we found only evidence for electrostatically induced proton release from peripheral amino acid residues but not from YZox itself. The positive charge stayed near YZox, and the rate of electron transfer was nearly independent of the pH. In core particles with an impaired Mn4 cluster, on the other hand, the rate of the electron transfer became strictly dependent on the protonation state of a single base (pK approximately 7). At pH < 7, the rate of electron transfer revealed the same slow rate (t1/2 approximately 35 microseconds) as that of proton release into the bulk. The deposition of a positive charge around YZox was no longer detected. A large H/D isotope effect (approximately 2.5) on these rates was also indicative of a steering of electron abstraction by proton transfer. That YZox was deprotonated into the bulk in inactive but not in oxygen-evolving material argues against the proposed role of YZox as an acceptor of hydrogen from water. Instead, the positive charge in its vicinity may shift the equilibrium from bound water to bound peroxide upon S3-->S4 as a prerequisite for the formation of oxygen upon S4-->S0.

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Kinetics of fluorescence changes on top of three different lipid bilayers due to lateral proton migration. The observation area was located at a distance of 70 μm from the area of proton release. Because all FPE molecules are surrounded by DPhPC or DPhPE molecules, they are anticipated to accept protons, which are released from these molecules. GMO does not possess ionizable moieties so that in case of two-dimensional diffusion, proton release from one FPE molecule seems to be required before the next FPE molecule may pick up the proton. Despite the huge differences in proton release rates from the different lipids, τ max for all three lipid bilayers was similar. That is, lateral proton diffusivity is independent of the choice of the lipid. The buffer contained 0.1 mM Capso (pH 9.0) and 100 mM NaCl. The F1 Wv/+ mice were then backcrossed to A/J mice for 10 generations, creating A/J N10 Wv/+ mice. These were then crossed to B6 Wv/+ mice to create F1 wildtype and F1 Wv/Wv mice for study. A/J mice had an increased airway resistance compared to B6 mice, and F1 mice had a naïve AHR phenotype equivalent to their parental A/J strain. F1 Wv/Wv mice displayed an airway resistance similar to normoresponsive B6 mice. Values represent mean ± SE, n = at least 10 in each group.

ATP synthase (F(O)F(1)) operates as two rotary motor/generators coupled by a common shaft. Both portions, F(1) and F(O), are rotary steppers. Their symmetries are mismatched (C(3) versus C(10-14)). We used the curvature of fluorescent actin filaments, attached to the rotating c-ring, as a spring balance (flexural rigidity of 8. 10(-26) Nm(2)) to gauge the angular profile of the output torque at F(O) during ATP hydrolysis by F(1) (see theoretical companion article (. Biophys. J. 81:1234-1244.)). The large average output torque (50 +/- 6 pN. nm) proved the absence of any slip. Variations of the torque were small, and the output free energy of the loaded enzyme decayed almost linearly over the angular reaction coordinate. Considering the threefold stepping and high activation barrier of the driving motor proper, the rather constant output torque implied a soft elastic power transmission between F(1) and F(O). It is considered as essential, not only for the robust operation of this ubiquitous enzyme under symmetry mismatch, but also for a high turnover rate of the two counteracting and stepping motor/generators.

Protonmotive force (the transmembrane difference in electrochemical potential of protons, ) drives ATP synthesis in bacteria, mitochondria, and chloroplasts. It has remained unsettled whether the entropic (chemical) component of relates to the difference in the proton activity between two bulk water phases (deltapH(B)) or between two membrane surfaces (deltapH(S)). To scrutinize whether deltapH(S) can deviate from deltapH(B), we modeled the behavior of protons at the membrane/water interface. We made use of the surprisingly low dielectric permittivity of interfacial water as determined by O. Teschke, G. Ceotto, and E. F. de Souza (O. Teschke, G. Ceotto, and E. F. de Sousa, 2001, PHYS: Rev. E. 64:011605). Electrostatic calculations revealed a potential barrier in the water phase some 0.5-1 nm away from the membrane surface. The barrier was higher for monovalent anions moving toward the surface (0.2-0.3 eV) than for monovalent cations (0.1-0.15 eV). By solving the Smoluchowski equation for protons spreading away from proton "pumps" at the surface, we found that the barrier could cause an elevation of the proton concentration at the interface. Taking typical values for the density of proton pumps and for their turnover rate, we calculated that a potential barrier of 0.12 eV yielded a steady-state pH(S) of approximately 6.0; the value of pH(S) was independent of pH in the bulk water phase under neutral and alkaline conditions. These results provide a rationale to solve the long-lasting problem of the seemingly insufficient protonmotive force in mesophilic and alkaliphilic bacteria.