Regulation of the F0F1-ATP synthase: The conformation of subunit ε might be determined by directionality of subunit γ rotation

AbstractF0F1-ATP synthase couples ATP synthesis/hydrolysis with transmembrane proton transport. The catalytic mechanism involves rotation of the γεc∼10-subunits complex relative to the rest of the enzyme.In the absence of protonmotive force the enzyme is inactivated by the tight binding of MgADP. Subunit ε also modulates the activity: its conformation can change from a contracted to extended form with C-terminus stretched towards F1. The latter form ihnibits ATP hydrolysis (but not synthesis).We propose that the directionality of the coiled-coil subunit γ rotation determines whether subunit ε is in contracted or extended form. Block of rotation by MgADP presumably induces the extended conformation of subunit ε. This conformation might serve as a safety lock, stabilizing the ADP-inhibited state upon de-energization and preventing spontaneous re-activation and wasteful ATP hydrolysis. The hypothesis merges the known regulatory effects of ADP, protonmotive force and conformational changes of subunit ε into a consistent picture

The role of subunit epsilon in the catalysis and regulation of FOF1-ATP synthase

AbstractThe regulation of ATP synthase activity is complex and involves several distinct mechanisms. In bacteria and chloroplasts, subunit epsilon plays an important role in this regulation, (i) affecting the efficiency of coupling, (ii) influencing the catalytic pathway, and (iii) selectively inhibiting ATP hydrolysis activity. Several experimental studies indicate that the regulation is achieved through large conformational transitions of the α-helical C-terminal domain of subunit epsilon that occur in response to membrane energization, change in ATP/ADP ratio or addition of inhibitors. This review summarizes the experimental data obtained on different organisms that clarify some basic features as well as some molecular details of this regulatory mechanism. Multiple functions of subunit epsilon, its role in the difference between the catalytic pathways of ATP synthesis and hydrolysis and its influence on the inhibition of ATP hydrolysis by ADP are also discussed

The cytochrome bc1 complex of Rhodobacter capsulatus: ubiquinol oxidation in a dimeric Q-cycle?

AbstractWe studied the cytochrome bc1 complex (hereafter bc) by flash excitation of Rhodobacter capsulatus chromatophores. The reduction of the high-potential heme bh of cytochrome b (at 561 nm) and of cytochromes c (at 552 nm) and the electrochromic absorption transients (at 524 nm) were monitored after the first and second flashes of light, respectively. We kept the ubiquinone pool oxidized in the dark and concerned for the ubiquinol formation in the photosynthetic reaction center only after the second flash. Surprisingly, the first flash caused the oxidation of about one ubiquinol per bc dimer. Based on these and other data we propose a dimeric Q-cycle where the energetically unfavorable oxidation of the first ubiquinol molecule by one of the bc monomers is driven by the energetically favorable oxidation of the second ubiquinol by the other bc monomer resulting in a pairwise oxidation of ubiquinol molecules by the dimeric bc in the dark. The residual unpaired ubiquinol supposedly remains on the enzyme and is then oxidized after the first flash

Low Dielectric Permittivity of Water at the Membrane Interface: Effect on the Energy Coupling Mechanism in Biological Membranes

Protonmotive force (the transmembrane difference in electrochemical potential of protons, [Formula: see text]) drives ATP synthesis in bacteria, mitochondria, and chloroplasts. It has remained unsettled whether the entropic (chemical) component of [Formula: see text] relates to the difference in the proton activity between two bulk water phases (ΔpH(B)) or between two membrane surfaces (ΔpH(S)). To scrutinize whether ΔpH(S) can deviate from ΔpH(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 ∼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

Mulkidjanian, Low dielectric permittivity of water at the membrane interface: effect on the energy coupling mechanism in biological membranes, Biophys

. 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 ;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

Conformational Transitions of Subunit ɛ in ATP Synthase from Thermophilic Bacillus PS3

Subunit ɛ of bacterial and chloroplast FOF1-ATP synthase is responsible for inhibition of ATPase activity. In Bacillus PS3 enzyme, subunit ɛ can adopt two conformations. In the “extended”, inhibitory conformation, its two C-terminal α-helices are stretched along subunit γ. In the “contracted”, noninhibitory conformation, these helices form a hairpin. The transition of subunit ɛ from an extended to a contracted state was studied in ATP synthase incorporated in Bacillus PS3 membranes at 59°C. Fluorescence energy resonance transfer between fluorophores introduced in the C-terminus of subunit ɛ and in the N-terminus of subunit γ was used to follow the conformational transition in real time. It was found that ATP induced the conformational transition from the extended to the contracted state (half-maximum transition extent at 140 μM ATP). ADP could neither prevent nor reverse the ATP-induced conformational change, but it did slow it down. Acid residues in the DELSEED region of subunit β were found to stabilize the extended conformation of ɛ. Binding of ATP directly to ɛ was not essential for the ATP-induced conformational change. The ATP concentration necessary for the half-maximal transition (140 μM) suggests that subunit ɛ probably adopts the extended state and strongly inhibits ATP hydrolysis only when the intracellular ATP level drops significantly below the normal value