Illuminating Voltage Sensor Paddling in Different Membrane Milieu.
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ABSTRACT:
SUBMITTER: Torbeev V
PROVIDER: S-EPMC7037485 | biostudies-literature | 2020 Feb
REPOSITORIES: biostudies-literature
Publications
Illuminating Voltage Sensor Paddling in Different Membrane Milieu.
Biophysical journal 20191001 4
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Project description:The interaction between membrane proteins and the surrounding membrane is becoming increasingly appreciated for its role in regulating protein function, protein localization, and membrane morphology. In particular, recent studies have suggested that membrane deformation is needed to stably accommodate proteins harboring charged amino acids in their transmembrane (TM) region, as it is energetically prohibitive to bury charge in the hydrophobic core of the bilayer. Unfortunately, current computational methods are poorly equipped for describing such deformations, as atomistic simulations are often too short to observe large-scale membrane reorganization and most continuum approaches assume a flat membrane. Previously, we developed a method that overcomes these shortcomings by using elasticity theory to characterize equilibrium membrane distortions in the presence of a TM protein, while using traditional continuum electrostatic and nonpolar energy models to determine the energy of the protein in the membrane. Here, we linked the elastostatics, electrostatics, and nonpolar numeric solvers to permit the calculation of energies for nontrivial membrane deformations. We then coupled this procedure to a robust search algorithm that identifies optimal membrane shapes for a TM protein of arbitrary chemical composition. This advance now permits us to explore a host of biological phenomena that were beyond the scope of our original method. We show that the energy required to embed charged residues in the membrane can be highly nonadditive, and our model provides a simple mechanical explanation for this nonadditivity. Our results also predict that isolated voltage sensor segments do not insert into rigid membranes, but membrane bending dramatically stabilizes these proteins in the bilayer despite their high charge content. Additionally, we use the model to explore hydrophobic mismatch with regard to nonpolar peptides and mechanosensitive channels. Our method is in quantitative agreement with molecular dynamics simulations at a tiny fraction of the computational cost.
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Project description:In humans, large conductance voltage- and calcium-dependent potassium (BK) channels are regulated allosterically by transmembrane voltage and intracellular Ca2+. Divalent cation binding sites reside within the gating ring formed by two Regulator of Conductance of Potassium (RCK) domains per subunit. Using patch-clamp fluorometry, we show that Ca2+ binding to the RCK1 domain triggers gating ring rearrangements that depend on transmembrane voltage. Because the gating ring is outside the electric field, this voltage sensitivity must originate from coupling to the voltage-dependent channel opening, the voltage sensor or both. Here we demonstrate that alterations of the voltage sensor, either by mutagenesis or regulation by auxiliary subunits, are paralleled by changes in the voltage dependence of the gating ring movements, whereas modifications of the relative open probability are not. These results strongly suggest that conformational changes of RCK1 domains are specifically coupled to the voltage sensor function during allosteric modulation of BK channels.
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Project description:Direct structural insights on the fundamental mechanisms of permeation, selectivity, and gating remain unavailable for the Na(+) and Ca(2+) channel families. Here, we report the spectroscopic structural characterization of the isolated Voltage-Sensor Domain (VSD) of the prokaryotic Na(+) channel NaChBac in a lipid bilayer. Site-directed spin-labeling and EPR spectroscopy were carried out for 118 mutants covering all of the VSD. EPR environmental data were used to unambiguously assign the secondary structure elements, define membrane insertion limits, and evaluate the activated conformation of the isolated-VSD in the membrane using restrain-driven molecular dynamics simulations. The overall three-dimensional fold of the NaChBac-VSD closely mirrors those seen in KvAP, Kv1.2, Kv1.2-2.1 chimera, and MlotiK1. However, in comparison to the membrane-embedded KvAP-VSD, the structural dynamics of the NaChBac-VSD reveals a much tighter helix packing, with subtle differences in the local environment of the gating charges and their interaction with the rest of the protein. Using cell complementation assays we show that the NaChBac-VSD can provide a conduit to the transport of ions in the resting or "down" conformation, a feature consistent with our EPR water accessibility measurements in the activated or "up" conformation. These results suggest that the overall architecture of VSD's is remarkably conserved among K(+) and Na(+) channels and that pathways for gating-pore currents may be intrinsic to most voltage-sensors. Cell complementation assays also provide information about the putative location of the gating charges in the "down/resting" state and hence a glimpse of the extent of conformational changes during activation.
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