scholarly journals Estimating the pKa values of basic and acidic side chains in ion channels using electrophysiological recordings: A robust approach to an elusive problem

2011 ◽  
Vol 79 (12) ◽  
pp. 3485-3493 ◽  
Author(s):  
Gisela D. Cymes ◽  
Claudio Grosman
Keyword(s):  
Ion Channels ◽  
Side Chains ◽  
Pka Values ◽  
Robust Approach ◽  
Electrophysiological Recordings

scholarly journals Identifying the elusive link between amino acid sequence and charge selectivity in pentameric ligand-gated ion channels

2016 ◽  
Vol 113 (45) ◽  
pp. E7106-E7115 ◽  
Author(s):  
Gisela D. Cymes ◽  
Claudio Grosman
Keyword(s):  
Ion Channels ◽  
Careful Analysis ◽  
Side Chains ◽  
Electrophysiological Recordings ◽  
Charge Selectivity ◽  
The Many ◽  
Backbone Atoms ◽  
Charge Sign ◽  
Gated Ion Channels ◽  
Ligand Gated Ion Channels

Among neurotransmitter-gated ion channels, the superfamily of pentameric ligand-gated ion channels (pLGICs) is unique in that its members display opposite permeant-ion charge selectivities despite sharing the same structural fold. Although much effort has been devoted to the identification of the mechanism underlying the cation-versus-anion selectivity of these channels, a careful analysis of past work reveals that discrepancies exist, that different explanations for the same phenomenon have often been put forth, and that no consensus view has yet been reached. To elucidate the molecular basis of charge selectivity for the superfamily as a whole, we performed extensive mutagenesis and electrophysiological recordings on six different cation-selective and anion-selective homologs from vertebrate, invertebrate, and bacterial origin. We present compelling evidence for the critical involvement of ionized side chains—whether pore-facing or buried—rather than backbone atoms and propose a mechanism whereby not only their charge sign but also their conformation determines charge selectivity. Insertions, deletions, and residue-to-residue mutations involving nonionizable residues in the intracellular end of the pore seem to affect charge selectivity by changing the rotamer preferences of the ionized side chains in the first turn of the M2 α-helices. We also found that, upon neutralization of the charged residues in the first turn of M2, the control of charge selectivity is handed over to the many other ionized side chains that decorate the pore. This explains the long-standing puzzle as to why the neutralization of the intracellular-mouth glutamates affects charge selectivity to markedly different extents in different cation-selective pLGICs.

scholarly journals Allosteric Modulation of Ligand Gated Ion Channels by Ivermectin

2014 ◽  
pp. S215-S224 ◽  
Author(s):  
H. ZEMKOVA ◽  
V. TVRDONOVA ◽  
A. BHATTACHARYA ◽  
M. JINDRICHOVA
Keyword(s):  
Ion Channels ◽  
Binding Site ◽  
Glycine Receptor ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Structural Determinants ◽  
P2x4 Receptor ◽  
Electrophysiological Recordings ◽  
Gated Ion Channels ◽  
Ligand Gated Ion Channels

Ivermectin acts as a positive allosteric regulator of several ligand-gated channels including the glutamate-gated chloride channel (GluCl),  aminobutyric acid type-A receptor, glycine receptor, neuronal α7-nicotinic receptor and purinergic P2X4 receptor. In most of the ivermectin-sensitive channels, the effects of ivermectin include the potentiation of agonist-induced currents at low concentrations and channel opening at higher concentrations. Based on mutagenesis, electrophysiological recordings and functional analysis of chimeras between ivermectin-sensitive and ivermectin-insensitive receptors, it has been concluded that ivermectin acts by insertion between transmembrane helices. The three-dimensional structure of C. elegans GluCl complexed with ivermectin has revealed the details of the ivermectin-binding site, however, no generic motif of amino acids could accurately predict ivermectin binding site for other ligand gated channels. Here, we will review what is currently known about ivermectin binding and modulation of Cys-loop receptor family of ligand-gated ion channels and what are the critical structural determinants underlying potentiation of the P2X4 receptor channel.

Protein

1995 ◽  
Author(s):  
Roland Lüthy ◽  
David Eisenberg
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Amino Acid ◽  
Isoelectric Point ◽  
Extinction Coefficient ◽  
Titration Curve ◽  
Side Chains ◽  
Pka Values ◽  
Extinction Coefficients ◽  
Molecular Weights

Given a protein sequence, the amino acid composition can be determined by counting the number of residues of each type. Then a molecular weight can be calculated by summing the molecular weights of the individual amino acid residues, taking into account the loss of one H2O molecule per peptide bond. Table 1 lists the molecular weights of the twenty amino acids and water. This approach assumes that the protein has not been covalently modified. Because of extensive glycosylation of some proteins, this approach can significantly underestimate the actual molecular weight. With the pKa values of Table 1, it is possible to calculate the theoretical charge of a protein at a given pH by summing the charges of the amino acid side chains and of the amino terminus and carboxyl terminus. By performing this calculation over a pH range, one obtains a theoretical titration curve and an isoelectric point (the pH at which the protein hasanetchargeof zero). This method assumes that all normally titratable groups are accessible to water, and that all side chains have the intrinsic pKa values listed in Table 1. This assumption is not completely correct, and consequently, the theoretical isoelectric point may differ from the experimentally determined value. Figure 1 shows the calculated titration curve for pancreatic ribonuclease: the calculated isoelectric point is 8.2, whereas the measured value is 9.6 (Lehninger, 1977). The calculation of extinction coefficients (Gill and von Hippel, 1989) is performed in much the same way as that of the isoelectric point Individual residues are treated as if they are free amino acids, and the overall extinction coefficient is calculated as the sum of the extinction coefficients of the residues. The same basic assumption is made: Residues are assumed to be in typical environments and not to show unusual absorption due to their local environments. In the case of the extinction coefficient, however, this assumption seems to be generally acceptable; calculated extinction coefficients are typically within a few percent of the experimentally determined value, and errors of more than 15% are rare (Gill and von Hippel, 1989).

Localizing the Charged Side Chains of Ion Channels within the Crowded Charge Models

2012 ◽  
Vol 9 (1) ◽  
pp. 766-773 ◽  
Author(s):  
Justin J. Finnerty ◽  
Robert Eisenberg ◽  
Paolo Carloni
Keyword(s):  
Ion Channels ◽  
Side Chains

scholarly journals Primary structure of peptides and ion channels. Role of amino acid side chains in voltage gating of melittin channels

1990 ◽  
Vol 58 (6) ◽  
pp. 1367-1375 ◽  
Author(s):  
M.T. Tosteson ◽  
O. Alvarez ◽  
W. Hubbell ◽  
R.M. Bieganski ◽  
C. Attenbach ◽  
...  
Keyword(s):  
Amino Acid ◽  
Ion Channels ◽  
Primary Structure ◽  
Voltage Gating ◽  
Side Chains ◽  
Amino Acid Side Chains

scholarly journals A flexible GAS belt responds to pore mutations changing the ion selectivity of proton-gated channels

2021 ◽  
Vol 154 (1) ◽  
Author(s):  
Zhuyuan Chen ◽  
Sheng Lin ◽  
Tianze Xie ◽  
Jin-Ming Lin ◽  
Cecilia M. Canessa
Keyword(s):  
Ion Channels ◽  
Pore Structure ◽  
Ion Selectivity ◽  
Hydration Shell ◽  
Postsynaptic Membrane ◽  
Membrane Depolarization ◽  
Side Chains ◽  
Open Conformation ◽  
Central Pore ◽  
Close Fit

Proton-gated ion channels conduct mainly Na+ to induce postsynaptic membrane depolarization. Finding the determinants of ion selectivity requires knowledge of the pore structure in the open conformation, but such information is not yet available. Here, the open conformation of the hASIC1a channel was computationally modeled, and functional effects of pore mutations were analyzed in light of the predicted structures. The open pore structure shows two constrictions of similar diameter formed by the backbone of the GAS belt and, right beneath it, by the side chains of H28 from the reentrant loop. Models of nonselective mutant channels, but not those that maintain ion selectivity, predict enlargement of the GAS belt, suggesting that this motif is quite flexible and that the loss of stabilizing interactions in the central pore leads to changes in size/shape of the belt. Our results are consistent with the “close-fit” mechanism governing selectivity of hASIC1a, wherein the backbone of the GAS substitutes at least part of the hydration shell of a permeant ion to enable crossing the pore constriction.

scholarly journals Electrophysiological Recordings on a Sounding Rocket: Report of a First Attempt Using Xenopus laevis Oocytes

2020 ◽  
Vol 5 (2) ◽  
pp. 43-56 ◽  
Author(s):  
Simon L. Wuest ◽  
Tobias Plüss ◽  
Christoph Hardegger ◽  
Mario Felder ◽  
Aaron Kunz ◽  
...  
Keyword(s):  
Ion Channels ◽  
Xenopus Laevis ◽  
Parabolic Flight ◽  
Sounding Rocket ◽  
Xenopus Laevis Oocytes ◽  
Patch Clamp Technique ◽  
Biological Responses ◽  
Electrophysiological Recordings ◽  
Electrophysiological Measurements ◽  
Microgravity Conditions

AbstractIt is not fully understood how cells detect external mechanical forces, but mechanosensitive ion channels play important roles in detecting and translating physical forces into biological responses (mechanotransduction). With the “OoClamp” device, we developed a tool to study electrophysiological processes, including the gating properties of ion channels under various gravity conditions. The “OoClamp” device uses an adapted patch clamp technique and is operational during parabolic flight and centrifugation up to 20 g. In the framework of the REXUS/BEXUS program, we have further developed the “OoClamp” device with the goal of conducting electrophysiological experiments aboard a flying sounding rocket. The aim of such an experiment was first to assess whether electrophysiological measurements of Xenopus laevis oocytes can be performed on sounding rocket flights, something that has never been done before. Second, we aimed to examine the gating properties of ion channels under microgravity conditions. The experiment was conducted in March 2016 on the REXUS 20 rocket. The post-flight analysis showed that all recording chambers were empty as the rocket reached the microgravity phase. A closer analysis of the flight data revealed that the oocytes were ripped apart a few seconds after the rocket launch. This first attempt at using sounding rockets as a research platform for electrophysiological recordings was therefore limited. Our modified “OoClamp” hardware was able to perform the necessary tasks for difficult electrophysiological recordings aboard a sounding rocket; however, the physical stresses during launch (acceleration and vibrations) did not support viability of Xenopus oocytes.

Metal bridges to probe membrane ion channel structure and function

2015 ◽  
Vol 6 (3) ◽  
pp. 191-203 ◽  
Author(s):  
Paul Linsdell
Keyword(s):  
Ion Channels ◽  
Ion Channel ◽  
Conformational Changes ◽  
Three Dimensional ◽  
Structure And Function ◽  
Side Chains ◽  
Channel Protein ◽  
Conformational Rearrangements ◽  
And Function ◽  
Different Parts

AbstractIon channels are integral membrane proteins that undergo important conformational changes as they open and close to control transmembrane flux of different ions. The molecular underpinnings of these dynamic conformational rearrangements are difficult to ascertain using current structural methods. Several functional approaches have been used to understand two- and three-dimensional dynamic structures of ion channels, based on the reactivity of the cysteine side-chain. Two-dimensional structural rearrangements, such as changes in the accessibility of different parts of the channel protein to the bulk solution on either side of the membrane, are used to define movements within the permeation pathway, such as those that open and close ion channel gates. Three-dimensional rearrangements – in which two different parts of the channel protein change their proximity during conformational changes – are probed by cross-linking or bridging together two cysteine side-chains. Particularly useful in this regard are so-called metal bridges formed when two or more cysteine side-chains form a high-affinity binding site for metal ions such as Cd2+ or Zn2+. This review describes the use of these different techniques for the study of ion channel dynamic structure and function, including a comprehensive review of the different kinds of conformational rearrangements that have been studied in different channel types via the identification of intra-molecular metal bridges. Factors that influence the affinities and conformational sensitivities of these metal bridges, as well as the kinds of structural inferences that can be drawn from these studies, are also discussed.

A highly stable membrane for vanadium flow batteries (VFBs) enabled by the selective degradation of ionic side chains

2022 ◽  
Author(s):  
Ziming Zhao ◽  
Qing Dai ◽  
Xiaofeng Li ◽  
Suobo Zhang ◽  
Shenghai Li ◽  
...  
Keyword(s):  
Ion Channels ◽  
Side Chains ◽  
Selective Degradation ◽  
Flow Batteries

The side chains detach from the main chains of polymers, which leads to a transition of ion channels to micropores.

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