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WT open and closed states A527L open and closed states A614G open and closed states W563L open and closed states W568L open and closed states A565L open and closed states The trajectories can be visualized using computational tools such as VMD or PyMol uploading the topology file (*.prmtop) and then the trajectory (*.dcd)., {"references":["https://doi.org/10.1038/s41467-023-36730-7"]}

The elusive activation/deactivation mechanism of hERG is investigated, a voltage-gated potassium channel involved in severe inherited and drug-induced cardiac channelopathies, including the Long QT Syndrome. Firstly, the available structural data are integrated by providing a homology model for the closed state of the channel. Secondly, molecular dynamics combined with a network analysis revealed two distinct pathways coupling the voltage sensor domain with the pore domain. Interestingly, some LQTS-related mutations known to impair the activation/deactivation mechanism are distributed along the identified pathways, which thus suggests a microscopic interpretation of their role. Split channels simulations clarify a surprising feature of this channel, which is still able to gate when a cut is introduced between the voltage sensor domain and the neighboring helix S5. In summary, the presented results suggest possible activation/deactivation mechanisms of non-domain-swapped potassium channels that may aid in biomedical applications.

In order to permeate a nanopore, an ion must overcome a dehydration energy barrier caused by the redistribution of surrounding water molecules. The redistribution is inhomogeneous, anisotropic and strongly position-dependent, resulting in complex patterns that are routinely observed in molecular dynamics simulations. Here, we study the physical origin of these patterns and of how they can be predicted and controlled. We introduce an analytic model able to predict the patterns in a graphene nanopore in terms of experimentally accessible radial distribution functions, giving results that agree well with molecular dynamics simulations. The patterns are attributable to a complex interplay of ionic hydration shells with water layers adjacent to the graphene membrane and with the hydration cloud of the nanopore rim atoms, and we discuss ways of controlling them. Our findings pave the way to designing required transport properties into nanoionic devices by optimising the structure of the hydration patterns.

Biological ion channels are fundamental to maintaining life. In this manuscript we apply our recently developed statistical and linear response theory to investigate Na+ conduction through the prokaryotic Na+ channel NaChBac. This work is extended theoretically by the derivation of ionic conductivity and current in an electrochemical gradient, thus enabling us to compare to a range of whole-cell data sets performed on this channel. Furthermore, we also compare the magnitudes of the currents and populations at each binding site to previously published single-channel recordings and molecular dynamics simulations respectively. In doing so, we find excellent agreement between theory and data, with predicted energy barriers at each of the four binding sites of ∼4,2.9,3.6, and 4kT.

We introduce a statistical and linear response theory of selective conduction in biological ion channels with multiple binding sites and possible point mutations. We derive an effective grand-canonical ensemble and generalised Einstein relations for the selectivity filter, assuming strongly coordinated ionic motion, and allowing for ionic Coulomb blockade. The theory agrees well with data from the KcsA K$^+$ channel and a mutant. We show that the Eisenman relations for thermodynamic selectivity follow from the condition for fast conduction and find that maximum conduction requires the binding sites to be nearly identical.

In order to permeate a nanopore, an ion must overcome a dehydration energy barrier caused by the redistribution of surrounding water molecules. The redistribution is inhomogeneous, anisotropic and strongly position-dependent, resulting in complex patterns that are routinely observed in molecular dynamics simulations. We now address the questions of the physical origin of these patterns and of how they can be predicted and controlled. We introduce an analytic model able to predict the patterns in terms of experimentally accessible radial distributions functions, yielding results that agree well with molecular dynamics simulations. We show that the patterns are attributable to a complex interplay of ionic hydration shells with water layers adjacent to the membrane and with the hydration cloud of the nanopore rim atoms, and we discuss ways of controlling them. Our findings pave the way to designing required transport properties into nanoionic devices by optimising the structure of the hydration patterns.

Voltage-gated sodium channels (NaVs) play fundamental roles in eukaryotes, but their exceptional size hinders their structural resolution. Bacterial NaVs are simplified homologues of their eukaryotic counterparts, but their use as models of eukaryotic Na+ channels is limited by their homotetrameric structure at odds with the asymmetric Selectivity Filter (SF) of eukaryotic NaVs. This work aims at mimicking the SF of eukaryotic NaVs by engineering radial asymmetry into the SF of bacterial channels. This goal was pursued with two approaches: the co-expression of different monomers of the NaChBac bacterial channel to induce the random assembly of heterotetramers, and the concatenation of four bacterial monomers to form a concatemer that can be targeted by site-specific mutagenesis. Patch-clamp measurements and Molecular Dynamics simulations showed that an additional gating charge in the SF leads to a significant increase in Na+ and a modest increase in the Ca2+ conductance in the NavMs concatemer in agreement with the behavior of the population of random heterotetramers with the highest proportion of channels with charge &minus, 5e. We thus showed that charge, despite being important, is not the only determinant of conduction and selectivity, and we created new tools extending the use of bacterial channels as models of eukaryotic counterparts.

Ion channel selectivity is essential for their function, yet the molecular basis of a channel's ability to select between ions is still rather controversial. In this work, using a combination of molecular dynamics simulations and electrophysiological current measurements we analyze the ability of the NaChBac channel to discriminate between calcium and sodium. Our simulations show that a single calcium ion can access the Selectivity Filter (SF) interacting so strongly with the glutamate ring so as to remain blocked inside. This is consistent with the tiny calcium currents recorded in our patch-clamp experiments. Two reasons explain this scenario. The first is the higher free energy of ion/SF binding of Ca(2+) with respect to Na(+). The second is the strong electrostatic repulsion exerted by the resident ion that turns back a second potentially incoming Ca(2+), preventing the knock-on permeation mechanism. Finally, we analyzed the possibility of the Anomalous Mole Fraction Effect (AMFE), i.e. the ability of micromolar Ca(2+) concentrations to block Na(+) currents. Current measurements in Na(+)/Ca(2+) mixed solutions excluded the AMFE, in agreement with metadynamics simulations showing the ability of a sodium ion to by-pass and partially displace the resident calcium. Our work supports a new scenario for Na(+)/Ca(2+) selectivity in the bacterial sodium channel, challenging the traditional notion of an exclusion mechanism strictly confining Ca(2+) ions outside the channel.

Selective permeation through graphene nanopores is attracting increasing interest as an efficient and cost-effective technique for water desalination and purification. In this work, using umbrella sampling and molecular dynamics simulations with constant electric field, we analyze the influence of pore charge on potassium and chloride ion permeation. As pore charge is increased, the barrier of the potential of mean force (PMF) gradually decreases until it turns into a well split in two subminima. While in the case of K+ this pattern can be explained as an increasing electrostatic compensation of the desolvation cost, in the case of Cl􀀀 the pattern can be attributed to the accumulation of a concentration polarization layer of potassium ions screening pore charge. Th analysis of potassium PMFs in terms of forces revealed a conflicting influence on permeation of van der Waals and electrostatic forces that both undergo an inversion of their direction as pore charge is increased. Even if the most important transition involves the interplay between the electrostatic forces exerted by graphene and water, the simulations also revealed an important role of the changing distribution of potassium and chloride ions. The influence of pore charge on the orientation of water molecules was also found to affect the van der Waals forces they exert on potassium.

We analyze selective ionic conduction through an artificial nanopore in a single graphene sheet, using molecular dynamics simulations and the prehistory probability distribution. We assess position-dependent changes in the number and orientation of water molecules in the first and second hydration shells of the ion as it crosses the nanopore. We reveal coupling between an ionic double layer near the sheet to the statistical properties of the hydration shells.

Selective permeation through graphene nanopores is attracting increasing interest as an efficient and cost-effective technique for water desalination and purification. In this work, using umbrella sampling and molecular dynamics simulations with constant electric field, we analyze the influence of pore charge on potassium and chloride ion permeation. As pore charge is increased, the barrier of the potential of mean force (PMF) gradually decreases until it turns into a well split in two subminima. While in the case of K

We consider the selective permeation of ions through narrow water-filled channels in the presence of strong interaction between the ions. These interactions lead to highly correlated ionic motion, which can conveniently be described via the concept of a quasiparticle. Here, we connect the quasiparticle’s effective potential and the multi-ion potential of the mean force, found through molecular dynamics simulations, and we validate the method on an analytical toy model of the KcsA channel. Possible future applications of the method to the connection between molecular dynamical calculations and the experimentally measured current-voltage and current-concentration characteristics of the channel are discussed.

We study the highly-concerted motion of ions in anarrow biological ion channel (KcsA) by considering the notionof a quasiparticle, with specific focus on the transition process.Namely, we show that the ion entering or exiting the channelis correlated with the position of the quasiparticle. This resultis of importance in the rate theories of ion conduction throughnarrow channels and artificial nanopores.

// Simona Reina 1,2* , Vanessa Checchetto 3,4,5,* , Rosaria Saletti 6 , Ankit Gupta 7,* , Deepti Chaturvedi 7,* , Carlo Guardiani 8 , Francesca Guarino 1,2 , Mariano Andrea Scorciapino 9 , Andrea Magri 1,2 , Salvatore Foti 6 , Matteo Ceccarelli 8,10,** , Angela Anna Messina 11,12,** , Radhakrishnan Mahalakshmi 7,** , Ildiko Szabo 3,4,** and Vito De Pinto 1,2 1 Department of Biomedicine and Biotechnology BIOMETEC, Section of Biology and Genetics, University of Catania, Catania, Italy 2 National Institute for Biomembranes and Biosystems, Section of Catania, Catania, Italy 3 Department of Biology, University of Padova, Padova, Italy 4 CNR Institute of Neurosciences, Padova, Italy 5 Department of Biomedical Sciences, University of Padova, Padova, Italy 6 Department of Chemical Sciences, Mass Spectrometry Unit, University of Catania, Catania, Italy 7 Molecular Biophysics Laboratory, Department of Biological Sciences, Indian Institute of Science Education and Research, Bhopal, India 8 Department of Physics, University of Cagliari, Cagliari, Italy 9 Department of Biomedical Sciences, Biochemistry Unit, University of Cagliari, Cagliari, Italy 10 Istituto Officina dei Materiali del Consiglio Nazionale delle Ricerche (IOM-CNR), UOS, Trieste, Italy 11 Department of Biological, Geological and Environmental Sciences, Section of Molecular Biology, University of Catania, Catania, Italy 12 National Institute for Biomembranes and Biosystems, Section of Catania, Catania, Italy * These authors have contributed equally to this work ** Co-corresponding author Correspondence to: Vito De Pinto, email: // Keywords : VDACs, cysteine oxidation, disulfide bridge, mitochondrial intermembrane space, mass spectrometry Received : October 21, 2015 Accepted : December 23, 2015 Published : January 08, 2016 Abstract Voltage-Dependent Anion selective Channels (VDAC) are pore-forming mitochondrial outer membrane proteins. In mammals VDAC3, the least characterized isoform, presents a set of cysteines predicted to be exposed toward the intermembrane space. We find that cysteines in VDAC3 can stay in different oxidation states. This was preliminary observed when, in our experimental conditions, completely lacking any reducing agent, VDAC3 presented a pattern of slightly different electrophoretic mobilities. This observation holds true both for rat liver mitochondrial VDAC3 and for recombinant and refolded human VDAC3. Mass spectroscopy revealed that cysteines 2 and 8 can form a disulfide bridge in native VDAC3. Single or combined site-directed mutagenesis of cysteines 2, 8 and 122 showed that the protein mobility in SDS-PAGE is influenced by the presence of cysteine and by the redox status. In addition, cysteines 2, 8 and 122 are involved in the stability control of the pore as shown by electrophysiology, complementation assays and chemico-physical characterization. Furthermore, a positive correlation between the pore conductance of the mutants and their ability to complement the growth of porin-less yeast mutant cells was found. Our work provides evidence for a complex oxidation pattern of a mitochondrial protein not directly involved in electron transport. The most likely biological meaning of this behavior is to buffer the ROS load and keep track of the redox level in the inter-membrane space, eventually signaling it through conformational changes.

The problem of predicting selective transport of ions through nano-pores from their structure in the biological and nano-technological systems is addressed. We use a molecular dynamics simulation to provide insight into the key physical parameters of nano-pores and develop a self-consistent analytic theory describing ionic conduction and selectivity through these devices. We analyse the ion's dehydration and excess chemical potential, derive an expression for the conductivity of the nano-pore, and emphasize the role of fluctuations in its performance. The theory is verified by comparison of the predicted current-voltage characteristics with the molecular dynamics results and experimental data obtained for a graphene nano-pore and the KcsA biological channel.