Critical Role of Water in the Binding of Volatile Anesthetics to Proteins

Wang, Hai Jing; Kleinhammes, Alfred; Tang, Pei; Xu, Yan; Wu, Yue

doi:10.1021/jp407115j

Cited by 11 publications

(22 citation statements)

References 47 publications

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“…We propose here that dewetting of hydrophobic cavities upon anesthetic binding is the primary determinant of affinity for propofol and sevoflurane for intersubunit sites on GABA A receptors. This result is consistent with a previous observation, using NMR, that a critical level of hydration was required for halothane to bind to proteins (65). To our knowledge, this is the first time the number of displaced water molecules has been correlated with affinity to explain GA specificity and selectivity.…”

Section: Resultssupporting

confidence: 93%

Competitive dewetting underlies site-specific binding of general anesthetics to GABA(A) receptors

Murlidaran

Hénin

Brannigan

2019

Preprint

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GABA(A) receptors are pentameric ligand-gated ion channels playing a critical role in the modulation of neuronal excitability. These inhibitory receptors, gated by -aminobutyric acid (GABA), can be potentiated and even directly activated by intravenous and inhalational anesthetics. Intersubunit cavities in the transmembrane domain have been consistently identified as putative binding sites by numerous experiment and simulation results. Synaptic GABA(A) receptors are predominantly found in a 2↵:2 :1 stoichiometry, with four unique inter-subunit interfaces. Experimental and computational results have suggested a perplexing specificity, given that cavity-lining residues are highly conserved, and the functional effects of general anesthetics are only weakly sensitive to most mutations of cavity residues. Here we use Molecular Dynamics simulations and thermodynamically rigorous alchemical free energy perturbation (AFEP) techniques to calculate affinities of the intravenous anesthetic propofol and the inhaled anesthetic sevoflurane to all intersubunit sites in a heteromeric GABA(A) receptor. We find that the best predictor of general anesthetic affinity for the intersubunit cavity sites is water displacement: combinations of anesthetic and binding site that displace more water molecules have higher affinities than those that displace fewer. The amount of water displacement is, in turn, a function of size of the general anesthetic, successful competition of the general anesthetic with water for the few hydrogen bonding partners in the site, and inaccessibility of the site to lipid acyl chains. The latter explains the surprisingly low affinity of GAs for the ↵ intersubunit site, which is missing a bulky methionine residue at the cavity entrance and can be occupied by acyl chains in the unbound state. Simulations also identify sevoflurane binding sites in the subunit centers and in the pore, but predict that these are lower affinity than the intersubunit sites. SignificanceAfter over a century of research, it is established that general anesthetics interact directly with hydrophobic cavities in proteins. We still do not know why not all small hydrophobic molecules can act as general anesthetics, or why not all hydrophobic cavities bind these molecules. General anesthetics can even select among homologous sites on one critical target, the GABA(A) heteropentamer, although the origins of selectivity are unknown. Here we used rigorous free energy calculations to find that binding affinity correlates with the number of released water molecules, which in turn depends upon the lipid content of the cavity without bound anesthetic. Results suggest a mechanism that reconciles lipid-centered and protein-centered theories, and which can directly inform design of new anesthetics.

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Section: Resultssupporting

confidence: 93%

Competitive dewetting underlies site-specific binding of general anesthetics to GABA(A) receptors

Murlidaran

Hénin

Brannigan

2019

Preprint

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“…Our groups’ recent work elucidated the binding mechanism of halothane and other typical general anesthetics 40 . Halothane cannot directly bind to the protein in the absence of hydration.…”

Section: Discussionmentioning

confidence: 99%

“…Further hydration of the protein has smaller effects on the enthalpic and entropic changes but still results in significant decrease in Gibbs free energy upon alcohol binding. A significant difference is recognized between the role of water in alcohol binding and its role in the binding of other general anesthetics such as halothane 40 . The present work revealed the importance of water-protein interactions in alcohol binding.…”

Section: Introductionmentioning

confidence: 99%

Dominant Alcohol–Protein Interaction via Hydration-Enabled Enthalpy-Driven Binding Mechanism

Chong

Kleinhammes

Tang³

et al. 2015

J. Phys. Chem. B

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Water plays an important role in weak associations of small drug molecules with proteins. Intense focus has been on binding-induced structural changes in the water network surrounding protein binding sites, especially their contributions to binding thermodynamics. However, water is also tightly coupled to protein conformations and dynamics, and so far little is known about the influence of water-protein interactions on ligand binding. Alcohols are a type of low-affinity drugs, and it remains unclear how water affects alcohol-protein interactions. Here, we present alcohol adsorption isotherms under controlled protein hydration using in-situ NMR detection. As functions of hydration level, Gibbs free energy, enthalpy, and entropy of binding were determined from the temperature dependence of isotherms. Two types of alcohol binding were found. The dominant type is low-affinity nonspecific binding, which is strongly dependent on temperature and the level of hydration. At low hydration levels, this nonspecific binding only occurs above a threshold of alcohol vapor pressure. An increased hydration level reduces this threshold, with it finally disappearing at a hydration level of h~0.2 (g water/g protein), gradually shifting alcohol binding from an entropy-driven to an enthalpy-driven process. Water at charged and polar groups on the protein surface was found to be particularly important in enabling this binding. Although further increase in hydration has smaller effects on the changes of binding enthalpy and entropy, it results in significant negative change in Gibbs free energy due to unmatched enthalpy-entropy compensation. These results show the crucial role of water-protein interplay in alcohol binding.

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“…Receptor mutation studies suggest that high-affinity propofol binding of GABA A receptors occurs at sites that are distinctly different from those that bind low-affinity volatile anesthetics, such as isoflurane [2] . Low-affinity binding sites are postulated to be amphipathic and water filled pockets within the receptor [3,4] . At least in the case of N-methyl-D-aspartate (NMDA) receptors, this low-affinity protein binding is subject to a cut-off effect associated with molar water solubility, but not molecular structure or size [5] .…”

Section: Introductionmentioning

confidence: 99%

GABA<sub>A</sub> Receptor Modulation by Phenyl Ring Compounds Is Associated with a Water Solubility Cut-Off Value

Brosnan

Pham²

2016

Pharmacology

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Background: The modulation of N-methyl-D-aspartate receptors is associated with a molar water solubility cut-off effect of approximately 1.1 mmol/l and hence are unaffected by significantly less soluble compounds. However, compounds with this molar water solubility are still able to modulate γ-aminobutyric acid type A (GABA_A) receptors. We hypothesized that GABA_A receptor modulation by phenolic compounds would exhibit cut-off at a molar water solubility value less than 1.1 mmol/l. Methods: GABA_A receptors consisting of human α₁ and rat β₂ and γ_2s subunits were expressed in Xenopus laevis oocytes, and drug responses were measured using standard 2-electrode voltage clamp techniques. Twenty substituted phenols and benzenes of similar size and molecular volume were studied at saturated aqueous concentrations. Reversible and statistically significant change in GABA_A receptor current that was 10% or greater in magnitude from the baseline response defined a positive drug effect. Results: All phenyl ring compounds with a molar water solubility value equal to or greater than 0.46 mmol/l positively modulated GABA_A receptor currents. No compounds with a molar water solubility value equal to or less than 0.10 mmol/l had any effect on GABA_A receptor currents. Saturated solutions of phenols with 2,6-dimethyl and 2,6-diisopropyl substituents also caused channel opening in the absence of GABA. Conclusions: The molar water solubility cut-off for GABA_A receptor modulation by phenyl ring compounds lies between 0.10 and 0.46 mmol/l. Data suggest that hydrocarbons, perhaps including inhaled anesthetics, might modulate GABA_A receptors by displacing water from one or more low-affinity amphipathic binding sites to induce conformational changes that increase ion conductance.

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Critical Role of Water in the Binding of Volatile Anesthetics to Proteins

Cited by 11 publications

References 47 publications

Competitive dewetting underlies site-specific binding of general anesthetics to GABA(A) receptors

Competitive dewetting underlies site-specific binding of general anesthetics to GABA(A) receptors

Dominant Alcohol–Protein Interaction via Hydration-Enabled Enthalpy-Driven Binding Mechanism

GABA<sub>A</sub> Receptor Modulation by Phenyl Ring Compounds Is Associated with a Water Solubility Cut-Off Value

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