Aqueous Accessibility to the Transmembrane Regions of Subunit c of the Escherichia coli F1F0 ATP Synthase

Abstract

sensitivity was restricted to a pocket of four residues lying directly behind Asp-61. Aqueous accessibility was also probed using Cd 2؉ , a membrane-impermeant soft metal ion with properties similar to Ag ؉ . Cd 2؉ inhibition was restricted to the I28C substitution in TMH1 and residues surrounding Asp-61 in TMH2. The overall pattern of inhibition, by all of the reagents tested, indicates highest accessibility on the cytoplasmic side of TMH2 and in a pocket of residues around Asp-61, including proximal residues in TMH1. Additionally subunit a was shown to mediate access to this region by the membrane-impermeant probe 2-(trimethylammonium)ethyl methanethiosulfonate. Based upon these results and other information, a pocket of aqueous accessible residues, bordered by the peripheral surface of TMH4 of subunit a, is proposed to extend from the cytoplasmic side of cTMH2 to Asp-61 in the center of the membrane.

Realistic simulations of the coupling between the protomotive force and the mechanical rotation of the F 0 -ATPase

“…Those who are familiar with water channels in proteins (34) may wonder how valid is our finding of the asymmetric PTR. We believe that this finding is quite robust since some of the a/c interfacial regions, in particular near the Asp-Arg pair, are clearly solvated, and the rest are likely to be accessible to water without major steric penalty (29). In fact, the overall results of the CG calculations appear not to be so sensitive to the exact cutoff value for the interfacial water region (tested for 6-8 Å).…”

“…The membrane block has a width of about 30 Å and the central Asp residues of the c-ring were placed almost at the middle of the membrane. Experimental evidences have suggested the presence of proton channels on the P and N side of the a/c interface that has access to the hydrophilic environment (28,29). In order to explore this effect in our CG model, we removed part of the membrane environment from the vicinity of the Asp-Arg pair in the interfacial regions.…”

Interacting cytoplasmic loops of subunits a and c of Escherichia coli F 1 F 0 ATP synthase gate H + transport to the cytoplasm

“…We previously reported an aqueous access pathway extending from the cytoplasm to the G58C in the center of cTMH2 that permitted access and modification by both small cations, such as Ag + and Cd +2 , and also larger reagents, such as NEM and MTSethyl-trimethylammonium (22,25). If such an aqueous pathway is to specifically conduct protons to the region of cGly58 and cAsp61, additional residues that specifically gate H + access or egress would be required at the cytoplasm surface.…”

“…The Ag + -sensitive substitutions on the periplasmic side of TMHs 2-5 cluster at the interior of the four-helix bundle predicted by cross-linking and could interact to form a continuous aqueous pathway extending from the periplasmic surface to the central region of the lipid bilayer (11,13,19,20). We have proposed that the movement of H + from the periplasmic half-channel and binding to the single ionized Asp61 in the c-ring is mediated by a swiveling of TMHs at the a-c subunit interface (16,(21)(22)(23)(24). This gating is thought to be coupled with ionization of a protonated cAsp61 in the adjacent subunit of the c-ring and with release of the H + into the cytoplasmic halfchannel at the subunit a-c interface.…”

“…This gating is thought to be coupled with ionization of a protonated cAsp61 in the adjacent subunit of the c-ring and with release of the H + into the cytoplasmic halfchannel at the subunit a-c interface. The route of aqueous access to the cytoplasmic side of the c subunit packing at the a-c interface has also been mapped by the chemical probing of Cys substitutions and, more recently, by molecular dynamics simulations (22,25,26).…”