A Cation-π Interaction in the Binding Site of the Glycine Receptor Is Mediated by a Phenylalanine Residue

Pless, Stephan A.; Millen, Kat S.; Hanek, Ariele P.; Lynch, Joseph W.; Lester, Henry A.; Lummis, Sarah C. R.; Dougherty, Dennis A.

doi:10.1523/jneurosci.2540-08.2008

Cited by 72 publications

(88 citation statements)

References 32 publications

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“…1B). Loop D contributes to the complementary (Ϫ) side of the GlyR-binding pocket (11,26). As shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

Magnitude of a Conformational Change in the Glycine Receptor β1-β2 Loop Is Correlated with Agonist Efficacy

Pless¹,

Lynch

2009

Journal of Biological Chemistry

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The efficacy of agonists at Cys-loop ion channel receptors is determined by the rate they isomerize receptors to a pre-open flip state. Once the flip state is reached, the shut-open reaction is similar for low and high efficacy agonists. The present study sought to identify a conformational change associated with the closed-flip transition in the ␣1-glycine receptor. We employed voltage-clamp fluorometry to compare ligand-binding domain conformational changes induced by the following agonists, listed from highest to lowest affinity and efficacy: glycine > ␤-alanine > taurine. Voltage-clamp fluorometry involves labeling introduced cysteines with environmentally sensitive fluorophores and inferring structural rearrangements from ligand-induced fluorescence changes. Agonist affinity and efficacy correlated inversely with maximum fluorescence magnitudes at labeled residues in ligand-binding domain loops D and E, suggesting that large conformational changes in this region preclude efficacious gating. However, agonist affinity and efficacy correlated directly with maximum fluorescence magnitudes from a label attached to A52C in loop 2, near the transmembrane domain interface. Because glycine experiences the largest affinity increase between closed and flip states, we propose that the magnitude of this fluorescence signal is directly proportional to the agonist affinity increase. In contrast, labeled residues in loops C, F, and the pre-M1 domain yielded agonist-independent fluorescence responses. Our results support the conclusion that a closed-flip conformation change, with a magnitude proportional to the agonist affinity increase from closed to flip states, occurs in the microenvironment of Ala-52. Glycine receptors (GlyRs)3 are pentameric chloride-selective ion channels that mediate fast inhibitory neurotransmission (1). They are members of the Cys-loop receptor family that includes the prototypical nicotinic acetylcholine receptor (nAChR), the ␥-aminobutyric acid type-A receptors (GABA A Rs), and serotonin type-3 receptors (5-HT 3 Rs). Recent structural studies have provided a wealth of information on the structure and function of this receptor family (2-6). In Cys-loop receptors, the ligand-binding domain (LBD) preceding the four transmembrane helices consists of two twisted ␤-sheets. The inner (vestibule facing) ␤-sheet comprises seven ␤-strands, while the outer ␤-sheet is formed by three ␤-strands (3). The ligand binding site is located at the interface of adjacent subunits and is lined by six domains: three loops from the principal and the complementary sides, termed A-C and D-F, respectively (3).GlyRs are activated by endogenous amino acid agonists in the following order of efficacy: glycine Ͼ ␤-alanine Ͼ taurine (7, 8). As these amino acids share considerable structural similarity (Fig. 1A), they are likely to compete for the same binding site (9 -11). A recent ground-breaking study on an intermediate pre-open state, the so-called "flip" state (12), has provided new insights into the mechanism of partial agon...

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“…1B). Loop D contributes to the complementary (Ϫ) side of the GlyR-binding pocket (11,26). As shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

Magnitude of a Conformational Change in the Glycine Receptor β1-β2 Loop Is Correlated with Agonist Efficacy

Pless¹,

Lynch

2009

Journal of Biological Chemistry

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“…Because Arg-65 forms a crucial component of the glycine-binding site (5,49), the nonconservative p.R65W mutation most likely disrupts glycine binding when functionally expressed with wild type subunits. Our finding that it is not trafficked to the surface when expressed as a homomer (Fig.…”

Section: Discussionmentioning

confidence: 99%

New Hyperekplexia Mutations Provide Insight into Glycine Receptor Assembly, Trafficking, and Activation Mechanisms

Bode

Wood

Mullins

et al. 2013

Journal of Biological Chemistry

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Background: Hyperekplexia mutations have provided much information about glycine receptor structure and function. Results: We identified and characterized nine new mutations. Dominant mutations resulted in spontaneous activation, whereas recessive mutations precluded surface expression. Conclusion: These data provide insight into glycine receptor activation mechanisms and surface expression determinants. Significance: The results enhance our understanding of hyperekplexia pathology and glycine receptor structure-function.

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“…A recent study using unnatural amino-acid derivatives of aromatic phenylalanine side chains has demonstrated that skeletal sodium channels form a cation-pi interaction with lidocaine and QX-314 at aromatic residue Phe1579 (Phe1760 in Na V 1.5), but not Tyr1586 (Tyr1767 in Na V 1.5) 25 . This class of electrostatic interaction occurs between a cation and the negative potential formed by the quadrupole moment on the face of an aromatic side chain 26,27 and has been shown to be responsible for ligand-receptor interactions in a variety of ion channels [28][29][30] . However, it remains to be shown whether such a cation-pi interaction is conserved in the cardiac sodium channel isoform and, more importantly, whether it is a general trait of all class I anti-arrhythmic drugs.…”

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confidence: 99%

Molecular basis for class Ib anti-arrhythmic inhibition of cardiac sodium channels

et al. 2011

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Cardiac sodium channels are established therapeutic targets for the management of inherited and acquired arrhythmias by class I anti-arrhythmic drugs (AADs). These drugs share a common target receptor bearing two highly conserved aromatic side chains, and are subdivided by the Vaughan-Williams classification system into classes Ia-c based on their distinct effects on the electrocardiogram. How can these drugs elicit distinct effects on the cardiac action potential by binding to a common receptor? Here we use fluorinated phenylalanine derivatives to test whether the electronegative surface potential of aromatic side chains contributes to inhibition by six class I AADs. Surprisingly, we find that class Ib AADs bind via a strong electrostatic cation-pi interaction, whereas class Ia and Ic AADs rely significantly less on this interaction. Our data shed new light on drug-target interactions underlying the inhibition of cardiac sodium channels by clinically relevant drugs and provide information for the directed design of AADs.

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A Cation-π Interaction in the Binding Site of the Glycine Receptor Is Mediated by a Phenylalanine Residue

Cited by 72 publications

References 32 publications

Magnitude of a Conformational Change in the Glycine Receptor β1-β2 Loop Is Correlated with Agonist Efficacy

Magnitude of a Conformational Change in the Glycine Receptor β1-β2 Loop Is Correlated with Agonist Efficacy

New Hyperekplexia Mutations Provide Insight into Glycine Receptor Assembly, Trafficking, and Activation Mechanisms

Molecular basis for class Ib anti-arrhythmic inhibition of cardiac sodium channels

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