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Nicotinic receptor interloop proline anchors beta1-beta2 and Cys loops in coupling agonist binding to channel gating.

Lee WY, Free CR, Sine SM - J. Gen. Physiol. (2008)

Bottom Line: We find that in contributing to channel gating, alphaVal 46 and alphaVal 132 of the signature Cys loop couple energetically to alphaPro 272.Furthermore, these residues are optimized in both their size and hydrophobicity to mediate rapid and efficient channel gating, suggesting naturally occurring substitutions at these positions enable a diverse range of gating rate constants among the Cys-loop receptor superfamily.The overall results indicate that alphaPro 272 functionally couples to flanking Val residues extending from the beta1-beta2 and Cys loops within the ACh binding to channel opening transduction pathway.

View Article: PubMed Central - PubMed

Affiliation: Receptor Biology Laboratory, Department of Physiology and Biomedical Engineering, Mayo Clinic College of Medicine, Rochester, MN 55905, USA.

ABSTRACT
Nicotinic acetylcholine receptors (AChRs) mediate rapid excitatory synaptic transmission throughout the peripheral and central nervous systems. They transduce binding of nerve-released ACh into opening of an intrinsic channel, yet the structural basis underlying transduction is not fully understood. Previous studies revealed a principal transduction pathway in which alphaArg 209 of the pre-M1 domain and alphaGlu 45 of the beta1-beta2 loop functionally link the two regions, positioning alphaVal 46 of the beta1-beta2 loop in a cavity formed by alphaPro 272 through alphaSer 269 of the M2-M3 loop. Here we investigate contributions of residues within and proximal to this pathway using single-channel kinetic analysis, site-directed mutagenesis, and thermodynamic mutant cycle analysis. We find that in contributing to channel gating, alphaVal 46 and alphaVal 132 of the signature Cys loop couple energetically to alphaPro 272. Furthermore, these residues are optimized in both their size and hydrophobicity to mediate rapid and efficient channel gating, suggesting naturally occurring substitutions at these positions enable a diverse range of gating rate constants among the Cys-loop receptor superfamily. The overall results indicate that alphaPro 272 functionally couples to flanking Val residues extending from the beta1-beta2 and Cys loops within the ACh binding to channel opening transduction pathway.

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Sequence alignment of subunits from the Cys-loop receptor superfamily. Secondary structural regions are shown at the top, with residue positions equivalent to αVal 46, αVal 132, and αPro 272 indicated by asterisks.
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fig2: Sequence alignment of subunits from the Cys-loop receptor superfamily. Secondary structural regions are shown at the top, with residue positions equivalent to αVal 46, αVal 132, and αPro 272 indicated by asterisks.

Mentions: The interface of the ligand binding and pore domains is depicted in the structural model of the Torpedo AChR α-subunit at a resolution of 4 Å (Fig. 1; PDB 2BG9; Unwin, 2005). Located within this interface is the principal coupling pathway that functionally links the two domains. The charge pair, αArg 209 and αGlu 45, is conserved across the Cys-loop superfamily in eukaryotes, but residues near this pair are not conserved. The position equivalent to αVal 46 of the β1–β2 loop can be substituted by Lys, Arg, Ala, Thr, or His; the position equivalent to αVal 132 of the Cys loop can be substituted by Ile, Leu, or Phe; and αPro 272 of the M2–M3 loop can be substituted by Lys or Thr (Fig. 2). The diversity of residues at the three positions suggests differences in the local structure flanking the principal coupling pathway contribute to the diverse range of channel gating kinetics among members of the Cys-loop receptor superfamily.


Nicotinic receptor interloop proline anchors beta1-beta2 and Cys loops in coupling agonist binding to channel gating.

Lee WY, Free CR, Sine SM - J. Gen. Physiol. (2008)

Sequence alignment of subunits from the Cys-loop receptor superfamily. Secondary structural regions are shown at the top, with residue positions equivalent to αVal 46, αVal 132, and αPro 272 indicated by asterisks.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2483337&req=5

fig2: Sequence alignment of subunits from the Cys-loop receptor superfamily. Secondary structural regions are shown at the top, with residue positions equivalent to αVal 46, αVal 132, and αPro 272 indicated by asterisks.
Mentions: The interface of the ligand binding and pore domains is depicted in the structural model of the Torpedo AChR α-subunit at a resolution of 4 Å (Fig. 1; PDB 2BG9; Unwin, 2005). Located within this interface is the principal coupling pathway that functionally links the two domains. The charge pair, αArg 209 and αGlu 45, is conserved across the Cys-loop superfamily in eukaryotes, but residues near this pair are not conserved. The position equivalent to αVal 46 of the β1–β2 loop can be substituted by Lys, Arg, Ala, Thr, or His; the position equivalent to αVal 132 of the Cys loop can be substituted by Ile, Leu, or Phe; and αPro 272 of the M2–M3 loop can be substituted by Lys or Thr (Fig. 2). The diversity of residues at the three positions suggests differences in the local structure flanking the principal coupling pathway contribute to the diverse range of channel gating kinetics among members of the Cys-loop receptor superfamily.

Bottom Line: We find that in contributing to channel gating, alphaVal 46 and alphaVal 132 of the signature Cys loop couple energetically to alphaPro 272.Furthermore, these residues are optimized in both their size and hydrophobicity to mediate rapid and efficient channel gating, suggesting naturally occurring substitutions at these positions enable a diverse range of gating rate constants among the Cys-loop receptor superfamily.The overall results indicate that alphaPro 272 functionally couples to flanking Val residues extending from the beta1-beta2 and Cys loops within the ACh binding to channel opening transduction pathway.

View Article: PubMed Central - PubMed

Affiliation: Receptor Biology Laboratory, Department of Physiology and Biomedical Engineering, Mayo Clinic College of Medicine, Rochester, MN 55905, USA.

ABSTRACT
Nicotinic acetylcholine receptors (AChRs) mediate rapid excitatory synaptic transmission throughout the peripheral and central nervous systems. They transduce binding of nerve-released ACh into opening of an intrinsic channel, yet the structural basis underlying transduction is not fully understood. Previous studies revealed a principal transduction pathway in which alphaArg 209 of the pre-M1 domain and alphaGlu 45 of the beta1-beta2 loop functionally link the two regions, positioning alphaVal 46 of the beta1-beta2 loop in a cavity formed by alphaPro 272 through alphaSer 269 of the M2-M3 loop. Here we investigate contributions of residues within and proximal to this pathway using single-channel kinetic analysis, site-directed mutagenesis, and thermodynamic mutant cycle analysis. We find that in contributing to channel gating, alphaVal 46 and alphaVal 132 of the signature Cys loop couple energetically to alphaPro 272. Furthermore, these residues are optimized in both their size and hydrophobicity to mediate rapid and efficient channel gating, suggesting naturally occurring substitutions at these positions enable a diverse range of gating rate constants among the Cys-loop receptor superfamily. The overall results indicate that alphaPro 272 functionally couples to flanking Val residues extending from the beta1-beta2 and Cys loops within the ACh binding to channel opening transduction pathway.

Show MeSH
Related in: MedlinePlus