An emerging consensus on voltage-dependent gating from computational modeling and molecular dynamics simulations
Ion Channel Gating
Kv1.2 Potassium Channel
Molecular Dynamics Simulation
Potassium Channels, Voltage-Gated
Voltage-Gated Sodium Channels
Permanent link to this recordhttp://hdl.handle.net/20.500.12613/5487
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AbstractDeveloping an understanding of the mechanism of voltage-gated ion channels in molecular terms requires knowledge of the structure of the active and resting conformations. Although the active-state conformation is known from x-ray structures, an atomic resolution structure of a voltage-dependent ion channel in the resting state is not currently available. This has motivated various efforts at using computational modeling methods and molecular dynamics (MD) simulations to provide the missing information. A comparison of recent computational results reveals an emerging consensus on voltage-dependent gating from computational modeling and MD simulations. This progress is highlighted in the broad context of preexisting work about voltage-gated channels. © 2012 Vargas et al.
Citation to related workRockefeller University Press
Has partJournal of General Physiology
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A MISSENSE MUTATION IN CONE PHOTORECEPTOR CYCLIC NUCLEOTIDE-GATED CHANNELS ASSOCIATED WITH CANINE DAYLIGHT BLINDNESS OFFERS INSIGHT INTO CHANNEL STRUCTURE AND FUNCTIONTanaka, Jacqueline; Gruberg, Edward R.; Chang, Frank N.; Carnevale, Vincenzo; O'Leary, Michael (Temple University. Libraries, 2013)Cone cyclic nucleotide-gated (CNG) channels are located in the retinal outer segments, mediating daylight color vision. The channel is a tetramer of A-type (CNGA3) and B-type (CNGB3) subunits. CNGA3 subunits are able to form homotetrameric channels, but CNGB3 exhibits channel function only when co-expressed with CNGA3. Mutations in the genes encoding these cone CNG subunits are associated with achromatopsia, an autosomal recessive genetic disorder which causes incomplete or complete loss of daylight and color vision. A missense mutation, aspartatic acid (Asp) to asparagine (Asn) at position 262 in the canine CNGB3 subunit (cB3-D262N), results in loss of cone function and therefore daylight blindness, highlighting the crucial role of this aspartic acid residue for proper channel biogenesis and/or function. Asp 262 is located in a conserved region of the second transmembrane segment containing three Asp residues designated the Tri-Asp motif. We exploit the conservation of these residues in CNGA3 subunits to examine the motif using a combination of experimental and computational approaches. Mutations of these conserved Asp residues result in a loss of nucleotide-activated currents and mislocalization in heterologous expression. Co-expressing CNGB3 Tri-Asp mutants with wild type CNGA3 results in functional channels, however, their electrophysiological characterization matches the properties of homomeric CNGA3 tetramers. This failure to record heteromeric currents implies that Asp/Asn mutations impact negatively both CNGA3 and CNGB3 subunits. A homology model of canine CNGA3 relaxed in a membrane using molecular dynamics simulations suggests that the Tri-Asp motif is involved in non-specific salt bridge pairings with positive residues of S3 - S4. We propose that the CNGB3-D262N mutation in daylight blind dogs results in the loss of these interactions and leads to an alteration of the electrostatic equilibrium in the S1 - S4 bundle. Because residues analogous to Tri-Asp residues in the voltage-gated Shaker K+ channel superfamily were implicated in monomer folding, we hypothesize that destabilizing these electrostatic interactions might impair the monomer folding state in D262N mutant CNG channels during biogenesis. Another missesnse sense mutation, Arginine (Arg) to tryptophan (Trp) at position 424 in the canine CNGA3 subunit (cA3-R424W), also results in loss of cone function. An amino acid sequence alignment with Shaker K+ channel superfamily indicates that this R424 residue is located in the C-terminal end of the sixth transmembrane segment. A3-R424W mutant channels resulted in no cyclic nucleotide-activated currents and mislocalization with intracellular aggregates. However, the localization of cA3-R424W mutant channels was not affected as severely as the Asp/Asn mutation in S2 Tri-Asp motif, showing a lot of cells with the proper localization of Golgi-like and membrane fluorescence. Moreover, the substitution of Arg 424 to Lysine (Lys), conserving the positive charge, preserved channel function in some cells, which is different from the results of the S2 Tri-Asp motif in which the Asp/Glu substitutions, conserving the negative charge, leads to loss of cyclic nucleotide-activated currents. Even though these missense mutations are both associated with canine daylight blindness, the Arg 424 residue might not be as critical for folding as the Tri-Asp residues in the S2 Tri-Asp motif and might be more of a problem in channel structure and function. The cA3 model relaxed with MD simulations indicated a possible interaction of Arg 424 with the Glu 304 residue in the S4-S5 linker. This hypothesis is supported by electrophysiological data in which the double mutation of reversing these residues, Glu 306 to Arg and Arg 424 to Glu (E306R-R424E) preserves channel function. In the model, this salt bridge appears to contribute to stabilization of the open pore state. The R424W mutation might disrupt the salt bridge formation, leading to deforming and closing the pore region.
A cyclic nucleotide-gated channel mutation associated with canine daylight blindness provides insight into a role for the S2 segment Tri-Asp motif in channel biogenesisTanaka, N; Delemotte, L; Klein, ML; Komáromy, AM; Tanaka, JC (2014-02-21)Cone cyclic nucleotide-gated channels are tetramers formed by CNGA3 and CNGB3 subunits; CNGA3 subunits function as homotetrameric channels but CNGB3 exhibits channel function only when co-expressed with CNGA3. An aspartatic acid (Asp) to asparagine (Asn) missense mutation at position 262 in the canine CNGB3 (D262N) subunit results in loss of cone function (daylight blindness), suggesting an important role for this aspartic acid residue in channel biogenesis and/or function. Asp 262 is located in a conserved region of the second transmembrane segment containing three Asp residues designated the Tri-Asp motif. This motif is conserved in all CNG channels. Here we examine mutations in canine CNGA3 homomeric channels using a combination of experimental and computational approaches. Mutations of these conserved Asp residues result in the absence of nucleotide-activated currents in heterologous expression. A fluorescent tag on CNGA3 shows mislocalization of mutant channels. Co-expressing CNGB3 Tri-Asp mutants with wild type CNGA3 results in some functional channels, however, their electrophysiological characterization matches the properties of homomeric CNGA3 channels. This failure to record heteromeric currents suggests that Asp/Asn mutations affect heteromeric subunit assembly. A homology model of S1-S6 of the CNGA3 channel was generated and relaxed in a membrane using molecular dynamics simulations. The model predicts that the Tri-Asp motif is involved in non-specific salt bridge pairings with positive residues of S3/S4. We propose that the D262N mutation in dogs with CNGB3-day blindness results in the loss of these inter-helical interactions altering the electrostatic equilibrium within in the S1-S4 bundle. Because residues analogous to Tri-Asp in the voltage-gated Shaker potassium channel family were implicated in monomer folding, we hypothesize that destabilizing these electrostatic interactions impairs the monomer folding state in D262N mutant CNG channels during biogenesis. © 2014 Tanaka et al.
Nucleoplasmic signals promote directed transmembrane protein import simultaneously via multiple channels of nuclear poresMudumbi, KC; Czapiewski, R; Ruba, A; Junod, SL; Li, Y; Luo, W; Ngo, C; Ospina, V; Schirmer, EC; Yang, W; Yang, Weidong|0000-0002-3554-3035 (2020-12-01)© 2020, The Author(s). Roughly 10% of eukaryotic transmembrane proteins are found on the nuclear membrane, yet how such proteins target and translocate to the nucleus remains in dispute. Most models propose transport through the nuclear pore complexes, but a central outstanding question is whether transit occurs through their central or peripheral channels. Using live-cell high-speed super-resolution single-molecule microscopy we could distinguish protein translocation through the central and peripheral channels, finding that most inner nuclear membrane proteins use only the peripheral channels, but some apparently extend intrinsically disordered domains containing nuclear localization signals into the central channel for directed nuclear transport. These nucleoplasmic signals are critical for central channel transport as their mutation blocks use of the central channels; however, the mutated proteins can still complete their translocation using only the peripheral channels, albeit at a reduced rate. Such proteins can still translocate using only the peripheral channels when central channel is blocked, but blocking the peripheral channels blocks translocation through both channels. This suggests that peripheral channel transport is the default mechanism that was adapted in evolution to include aspects of receptor-mediated central channel transport for directed trafficking of certain membrane proteins.