Activation of acetylcholine receptor channels by covalently bound agonists in cultured rat myoballs.

Chabala, L D; Lester, Henry A.

doi:10.1113/jphysiol.1986.sp016242

Cited by 38 publications

(25 citation statements)

References 46 publications

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“…This model suggests that any means of positioning a quaternary ammonium in close proximity to ␣Trp-149 might produce an active receptor. In classic experiments, tethered agonists were obtained by attaching structures containing quaternary ammonium groups to the cysteines produced by reducing the highly conserved Cys-192-Cys-193 disulfide of the ␣ subunit, leading to the conclusion that the quaternary ammonium group interacts with a moiety within 9 Å of the disulfide (30,31). We applied a similar strategy at position ␣149, incorporating the unnatural amino acid Tyr-O3Q (Tyr-O-(CH 2 ) 3 -N(CH 3 ) 3 ϩ , see Fig.…”

Section: Resultsmentioning

confidence: 99%

From ab initio quantum mechanics to molecular neurobiology: A cation–π binding site in the nicotinic receptor

Zhong

Gallivan

Zhang

et al. 1998

Proc. Natl. Acad. Sci. U.S.A.

553

505

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The nicotinic acetylcholine receptor is the prototype ligand-gated ion channel. A number of aromatic amino acids have been identified as contributing to the agonist binding site, suggesting that cation-interactions may be involved in binding the quaternary ammonium group of the agonist, acetylcholine. Here we show a compelling correlation between: (i) ab initio quantum mechanical predictions of cation-binding abilities and (ii) EC 50 values for acetylcholine at the receptor for a series of tryptophan derivatives that were incorporated into the receptor by using the in vivo nonsense-suppression method for unnatural amino acid incorporation. Such a correlation is seen at one, and only one, of the aromatic residues-tryptophan-149 of the ␣ subunit. This finding indicates that, on binding, the cationic, quaternary ammonium group of acetylcholine makes van der Waals contact with the indole side chain of ␣ tryptophan-149, providing the most precise structural information to date on this receptor. Consistent with this model, a tethered quaternary ammonium group emanating from position ␣149 produces a constitutively active receptor.

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Section: Resultsmentioning

confidence: 99%

From ab initio quantum mechanics to molecular neurobiology: A cation–π binding site in the nicotinic receptor

Zhong

Gallivan

Zhang

et al. 1998

Proc. Natl. Acad. Sci. U.S.A.

553

505

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“…Early studies with native nAChRs established that bromoacetylcholine acted as a covalent agonist after reduction of the ␣Cys-192/193 disulfide (Damle and Karlin, 1978;Chabala and Lester, 1986). We established previously that covalent modification of ␣Y198C with MTSET, MTSPT, or bromoacetylcholine resulted in the addition of a covalent agonist (Sullivan and Cohen, 2000).…”

Section: Discussionmentioning

confidence: 99%

Mapping the Agonist Binding Site of the Nicotinic Acetylcholine Receptor by Cysteine Scanning Mutagenesis: Antagonist Footprint and Secondary Structure Prediction

2002

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To further define the surface of the Torpedo californica nicotinic acetylcholine receptor (nAChR) contributing to the agonist binding site structure, we used the substituted Cys accessibility method to identify novel residues and determined the "footprint" of residues protected from modification by the reversible competitive antagonist d-tubocurarine (dTC). nAChRs containing single Cys substitutions within regions of the ␣-or ␥-subunit primary structure known to contribute to the agonist binding site were expressed in Xenopus laevis oocytes. Cys substitutions in binding site segments A (␣Tyr-93 and ␣Asn-94), C (␣Tyr-198), and D (␥Glu-57) had been shown previously to be accessible for modification. We now introduced cysteines from ␣Asp-195 to ␣Ile-201 and from ␥Ala-106 to ␥Asp-113 and identified positions accessible for modification in segments C ) and E (␥Asn-107 and ␥Leu-109). dTC protected against alkylation in segments D (␥Glu-57) and E (␥Leu-109) but not in segment A (␣Tyr-93 and ␣Asn-94). In segment C, dTC protection experiments revealed a pattern in which every other residue (␣196, ␣198, and ␣200, but not ␣197 or ␣201) was protected from alkylation. This pattern of protection provides evidence that bound dTC is near amino acids in segments C, D, and E but not in segment A, and identifies a ␤-strand surface within segment C contributing to the binding site. These results are discussed in terms of a homology model, based on the molluscan acetylcholine binding protein crystal structure, of the T. californica nAChR agonist binding site.

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“…PTLs have been used in ion channels to conditionally present an agonist to an allosteric regulatory site, originally in the nicotinic acetylcholine receptor (7)(8)(9)(10), and recently in a kainate receptor, iGluR6 (LiGluR) (11)(12)(13) or to conditionally present a blocker to the Shaker K ϩ channel (SPARK) (14,15). PTLs contain the ligand (agonist/antagonist or blocker) at one end, a reactive group that attaches covalently to the protein at the other end, and a linker in the middle that contains a photoisomerizable moieity, such as azobenzene.…”

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confidence: 99%

Nanosculpting reversed wavelength sensitivity into a photoswitchable iGluR

Szobota

Lau

Gorostiza

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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Photoswitched tethered ligands (PTLs) can be used to remotely control protein function with light. We have studied the geometric and conformational factors that determine the efficacy of PTL gating in the ionotropic glutamate receptor iGluR6 using a family of photoiosomerizable MAG (maleimide-azobenzene-glutamate) PTLs that covalently attach to the clamshell ligand-binding domain. Experiments and molecular dynamics simulations of the modified proteins show that optical switching depends on 2 factors: (i) the relative occupancy of the binding pocket in the 2 photoisomers of MAG and (ii) the degree of clamshell closure that is possible given the disposition of the MAG linker. A synthesized short version of MAG turns the channel on in either the cis or trans state, depending on the point of attachment. This yin/yang optical control makes it possible for 1 wavelength of light to elicit action potentials in one set of neurons, while deexciting a second set of neurons in the same preparation, whereas a second wavelength has the opposite effect. The ability to generate opposite responses with a single PTL and 2 versions of a target channel, which can be expressed in different cell types, paves the way for engineering opponency in neurons that mediate opposing functions.glutamate receptor ͉ ion channel ͉ optics ͉ photoswitch

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Activation of acetylcholine receptor channels by covalently bound agonists in cultured rat myoballs.

Cited by 38 publications

References 46 publications

From ab initio quantum mechanics to molecular neurobiology: A cation–π binding site in the nicotinic receptor

From ab initio quantum mechanics to molecular neurobiology: A cation–π binding site in the nicotinic receptor

Mapping the Agonist Binding Site of the Nicotinic Acetylcholine Receptor by Cysteine Scanning Mutagenesis: Antagonist Footprint and Secondary Structure Prediction

Nanosculpting reversed wavelength sensitivity into a photoswitchable iGluR

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