Interactions Between a Voltage Sensor and a Toxin via Multiscale Simulations

Gating-modifier toxins inhibit voltage-gated ion channels by binding the voltage sensors (VS) and altering the energetics of voltage-dependent gating. These toxins are thought to gain access to the VS via the membrane (i.e., by partitioning from water into the membrane before binding the VS). We used serial multiscale molecular-dynamics (MD) simulations, via a combination of coarse-grained (CG) and atomistic (AT) simulations, to study how the toxin VSTx1, which inhibits the archeabacterial voltage-gated potassium channel KvAP, interacts with an isolated membrane-embedded VS domain. In the CG simulations, VSTx1, which was initially located in water, partitioned into the headgroup/water interface of the lipid bilayer before binding the VS. The CG configurations were used to generate AT representations of the system, which were subjected to AT-MD to further evaluate the stability of the complex and refine the predicted VS/toxin interface. VSTx1 interacted with a binding site on the VS formed by the C-terminus of S1, the S1-S2 linker, and the N-terminus of S4. The predicted VS/toxin interactions are suggestive of toxin-mediated perturbations of the interaction between the VS and the pore domain of Kv channels, and of the membrane. Our simulations support a membrane-access mechanism of inhibition of Kv channels by VS toxins. Overall, the results show that serial multiscale MD simulations may be used to model a two-stage process of protein-bilayer and protein-protein interactions within a membrane

Voltage sensors, toxins & membranes : a simulation study

Molecular dynamics (MO) simulations were used to study the interaction of voltage-gated potassium (Kv) channel toxins with lipid bilayers. The toxins studied were YSTx1 (Yoltage Sensor Toxin 1) and SGTx1 (Seodm griseipes Toxin 1), gating-modifier toxins from tarantula venom that bind to the voltage sensors (YS) of Kv channels and inhibit the channels by altering the energetics of voltagedependent gating. The mechanism of interaction and the depth of binding of these toxins with lipid bilayers is of interest in the context of structural models of voltage-dependent gating in Kv channels. In particular, characterizing the interaction of these toxins with lipid bilayers provided insights into the structure and dynamics of the YS of Kv channels. Atomistic (AT) MO simulations were used to localize YSTx1 and SGTx1 in lipid bilayers. Both toxins preferred a location close to the ·membrane/water interface consistent with their amphipathic molecular surface. The nature of the interactions that stabilized the toxins in the membrane was investigated. ExtendedMOsimulations with a coarse-grained (CG) protein and lipid model revealed dynamic toxin partitioning of SGTx1 from bulk water to the membrane/water interface. CG MO simulations with a total simulation time of > 30 ps were used to estimate the 10 potential of mean force (PMF) profile of YSTx1 along the bilayer normal of lipid bilayers. The PMF profiles suggest it is energetically favorable for YSTx1 to partition from bulk water to the membrane/water interface. The bilayer deformed as it interacted with the toxin, and bilayer deformation influenced the PMF profiles. AT MO simulations with a total simulation time of > 1.6 ps were also used to estimate the 10 PMF profile of YSTx1 along the bilayer normal. A novel approach was taken whereby information derived from the CG MO simulations were used to initialize the AT MO simulations. Comparisons were· made between the CG and AT PMF profiles, and that derived from an implicit membrane/solvent model. CG MO simulations were used to estimate an angular PMF profile of a 54 helix (which carries the gating charges in the YS of Kv channels) in lipid bilayers. The CG PMF profiles suggested that it is overall energetically favorable for 54 to adopt a membrane surface orientation compared to a transmembrane (TM) orientation. Interestingly, 54 could not be (meta) stably inserted in a TM orientation in membranes with reduced concentration of negatively-charged lipid phosphate moieties. Finally, CG MO simulations were performed to probe the interaction of a membrane-embedded YS with YSTx1.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Membrane Insertion of a Voltage Sensor Helix

Most membrane proteins contain a transmembrane (TM) domain made up of a bundle of lipid-bilayer-spanning α-helices. TM α-helices are generally composed of a core of largely hydrophobic amino acids, with basic and aromatic amino acids at each end of the helix forming interactions with the lipid headgroups and water. In contrast, the S4 helix of ion channel voltage sensor (VS) domains contains four or five basic (largely arginine) side chains along its length and yet adopts a TM orientation as part of an independently stable VS domain. Multiscale molecular dynamics simulations are used to explore how a charged TM S4 α-helix may be stabilized in a lipid bilayer, which is of relevance in the context of mechanisms of translocon-mediated insertion of S4. Free-energy profiles for insertion of the S4 helix into a phospholipid bilayer suggest that it is thermodynamically favorable for S4 to insert from water to the center of the membrane, where the helix adopts a TM orientation. This is consistent with crystal structures of Kv channels, biophysical studies of isolated VS domains in lipid bilayers, and studies of translocon-mediated S4 helix insertion. Decomposition of the free-energy profiles reveals the underlying physical basis for TM stability, whereby the preference of the hydrophobic residues of S4 to enter the bilayer dominates over the free-energy penalty for inserting charged residues, accompanied by local distortion of the bilayer and penetration of waters. We show that the unique combination of charged and hydrophobic residues in S4 allows it to insert stably into the membrane

The Energetics of Transmembrane Helix Insertion into a Lipid Bilayer

Free energy profiles for insertion of a hydrophobic transmembrane protein α-helix (M2 from CFTR) into a lipid bilayer have been calculated using coarse-grained molecular dynamics simulations and umbrella sampling to yield potentials of mean force along a reaction path corresponding to translation of a helix across a lipid bilayer. The calculated free energy of insertion is smaller when a bilayer with a thinner hydrophobic region is used. The free energies of insertion from the potentials of mean force are compared with those derived from a number of hydrophobicity scales and with those derived from translocon-mediated insertion. This comparison supports recent models of translocon-mediated insertion and in particular suggests that: 1), helices in an about-to-be-inserted state may be located in a hydrophobic region somewhat thinner than the core of a lipid bilayer; and/or 2), helices in a not-to-be-inserted state may experience an environment more akin (e.g., in polarity/hydrophobicity) to the bilayer/water interface than to bulk water

Lipid Bilayer Deformation and the Free Energy of Interaction of a Kv Channel Gating-Modifier Toxin

A number of membrane proteins act via binding at the water/lipid bilayer interface. An important example of such proteins is provided by the gating-modifier toxins that act on voltage-gated potassium (Kv) channels. They are thought to partition to the headgroup region of lipid bilayers, and so provide a good system for probing the nature of interactions of a protein with the water/bilayer interface. We used coarse-grained molecular dynamics simulations to compute the one-dimensional potential of mean force (i.e., free energy) profile that governs the interaction between a Kv channel gating-modifier toxin (VSTx1) and model phospholipid bilayers. The reaction coordinate sampled corresponds to the position of the toxin along the bilayer normal. The course-grained representation of the protein and lipids enabled us to explore extended time periods, revealing aspects of toxin/bilayer dynamics and energetics that would be difficult to observe on the timescales currently afforded by atomistic molecular dynamics simulations. In particular, we show for this model system that the bilayer deforms as it interacts with the toxin, and that such deformations perturb the free energy profile. Bilayer deformation therefore adds an additional layer of complexity to be addressed in investigations of membrane/protein systems. In particular, one should allow for local deformations that may arise due to the spatial array of charged and hydrophobic elements of an interfacially located membrane protein

The Interaction of Phospholipase A2 with a Phospholipid Bilayer: Coarse-Grained Molecular Dynamics Simulations

A number of membrane-active enzymes act in a complex environment formed by the interface between a lipid bilayer and bulk water. Although x-ray diffraction studies yield structures of isolated enzyme molecules, a detailed characterization of their interactions with the interface requires a measure of how deeply such a membrane-associated protein penetrates into a lipid bilayer. Here, we apply coarse-grained (CG) molecular dynamics (MD) simulations to probe the interaction of porcine pancreatic phospholipase A2 (PLA2) with a lipid bilayer containing palmitoyl-oleoyl-phosphatidyl choline and palmitoyl-oleoyl-phosphatidyl glycerol molecules. We also used a configuration from a CG-MD trajectory to initiate two atomistic (AT) MD simulations. The results of the CG and AT simulations are evaluated by comparison with available experimental data. The membrane-binding surface of PLA2 consists of a patch of hydrophobic residues surrounded by polar and basic residues. We show this proposed footprint interacts preferentially with the anionic headgroups of the palmitoyl-oleoyl-phosphatidyl glycerol molecules. Thus, both electrostatic and hydrophobic interactions determine the location of PLA2 relative to the bilayer. From a general perspective, this study demonstrates that CG-MD simulations may be used to reveal the orientation and location of a membrane-surface-bound protein relative to a lipid bilayer, which may subsequently be refined by AT-MD simulations to probe more detailed interactions