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Thermodynamics and mechanism of the interaction of willardiine partial agonists with a glutamate receptor: implications for drug development.

Martinez M, Ahmed AH, Loh AP, Oswald RE - Biochemistry (2014)

Bottom Line: The binding of the charged form is largely driven by enthalpy, while that of the uncharged form is largely driven by entropy.This is due at least in part to changes in the hydrogen bonding network within the binding site involving one water molecule.This work illustrates the importance of charge to the thermodynamics of binding of agonists and antagonists to AMPA receptors and provides clues for further drug discovery.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Medicine, Cornell University , Ithaca, New York 14853, United States.

ABSTRACT
Understanding the thermodynamics of binding of a lead compound to a receptor can provide valuable information for drug design. The binding of compounds, particularly partial agonists, to subtypes of the α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptor is, in some cases, driven by increases in entropy. Using a series of partial agonists based on the structure of the natural product, willardiine, we show that the charged state of the ligand determines the enthalpic contribution to binding. Willardiines have uracil rings with pKa values ranging from 5.5 to 10. The binding of the charged form is largely driven by enthalpy, while that of the uncharged form is largely driven by entropy. This is due at least in part to changes in the hydrogen bonding network within the binding site involving one water molecule. This work illustrates the importance of charge to the thermodynamics of binding of agonists and antagonists to AMPA receptors and provides clues for further drug discovery.

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(A) Ionization statesof the uracil ring of NW. Similar ionizationstates are possible for the other willardiine derivatives. (B) Effectof a change in pH on the thermodynamic parameters for NW (left) andHW (right) at 10 °C. For both compounds, the pH at which theionized state is favored is shown on the left and that for which theun-ionized state is favored is shown on the right. (C) Structure ofthe binding site, showing the direct interactions between NW and theGluA2 LBD [Protein Data Bank (PDB) entry 3RTW(32)]. (D) Structureof the binding site for NW showing the hydrogen bonding network associatedwith the nitrogen at position 3 of the willardiine ring in the chargedform (pH 6.5, PDB entry 3RTW(32)). On the left is thestructure and on the right a schematic. In the schematic, NW is coloredblue, the protein black, and water green and the H-bonds are coloredred. (E) Structure of NW bound to the GluA2 LBD obtained at pH 3.5(PDB entry 4Q30). At this pH, the ring is largely uncharged. The formatting of thispanel is similar to that of panel D. Note the change in the H-bondingnetwork. The potential H-bond between the hydroxyl of S654 and theuracil carbonyl is shown with a question mark because the distancebetween the two oxygens is relatively long, 3.5 Å, and the anglebetween this H-bond and the H-bond with the water is not favorable(80°).
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fig2: (A) Ionization statesof the uracil ring of NW. Similar ionizationstates are possible for the other willardiine derivatives. (B) Effectof a change in pH on the thermodynamic parameters for NW (left) andHW (right) at 10 °C. For both compounds, the pH at which theionized state is favored is shown on the left and that for which theun-ionized state is favored is shown on the right. (C) Structure ofthe binding site, showing the direct interactions between NW and theGluA2 LBD [Protein Data Bank (PDB) entry 3RTW(32)]. (D) Structureof the binding site for NW showing the hydrogen bonding network associatedwith the nitrogen at position 3 of the willardiine ring in the chargedform (pH 6.5, PDB entry 3RTW(32)). On the left is thestructure and on the right a schematic. In the schematic, NW is coloredblue, the protein black, and water green and the H-bonds are coloredred. (E) Structure of NW bound to the GluA2 LBD obtained at pH 3.5(PDB entry 4Q30). At this pH, the ring is largely uncharged. The formatting of thispanel is similar to that of panel D. Note the change in the H-bondingnetwork. The potential H-bond between the hydroxyl of S654 and theuracil carbonyl is shown with a question mark because the distancebetween the two oxygens is relatively long, 3.5 Å, and the anglebetween this H-bond and the H-bond with the water is not favorable(80°).

Mentions: The influence ofsubstituent electronegativity on the thermodynamicparameters suggested that the differing pKa of the uracil ring, due to the different analogues in position 5,may affect the thermodynamics of binding. While the pKa of all the halogenated uracils is ∼8, those ofHW and NW are approximately 10 and 6, respectively.29,30 This means that willardiine and the halogenated analogues were,at physiological pH, in mostly uncharged (protonated) states (Figure 2A). NW, however, was in a mostly charged (deprotonated)state. The GluA2 LBD is well-behaved between pH 4 and 10, with minorchanges in the NMR 1H–15N HSQC spectrum.9 We tested the possibility that the charged stateof the ligand could affect the enthalpy versus entropy distributionby changing the pH. Thus, binding of largely uncharged (protonateduracil ring) NW was examined at pH 4, and binding of largely charged(deprotonated uracil ring) HW was examined at pH 10.


Thermodynamics and mechanism of the interaction of willardiine partial agonists with a glutamate receptor: implications for drug development.

Martinez M, Ahmed AH, Loh AP, Oswald RE - Biochemistry (2014)

(A) Ionization statesof the uracil ring of NW. Similar ionizationstates are possible for the other willardiine derivatives. (B) Effectof a change in pH on the thermodynamic parameters for NW (left) andHW (right) at 10 °C. For both compounds, the pH at which theionized state is favored is shown on the left and that for which theun-ionized state is favored is shown on the right. (C) Structure ofthe binding site, showing the direct interactions between NW and theGluA2 LBD [Protein Data Bank (PDB) entry 3RTW(32)]. (D) Structureof the binding site for NW showing the hydrogen bonding network associatedwith the nitrogen at position 3 of the willardiine ring in the chargedform (pH 6.5, PDB entry 3RTW(32)). On the left is thestructure and on the right a schematic. In the schematic, NW is coloredblue, the protein black, and water green and the H-bonds are coloredred. (E) Structure of NW bound to the GluA2 LBD obtained at pH 3.5(PDB entry 4Q30). At this pH, the ring is largely uncharged. The formatting of thispanel is similar to that of panel D. Note the change in the H-bondingnetwork. The potential H-bond between the hydroxyl of S654 and theuracil carbonyl is shown with a question mark because the distancebetween the two oxygens is relatively long, 3.5 Å, and the anglebetween this H-bond and the H-bond with the water is not favorable(80°).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4215890&req=5

fig2: (A) Ionization statesof the uracil ring of NW. Similar ionizationstates are possible for the other willardiine derivatives. (B) Effectof a change in pH on the thermodynamic parameters for NW (left) andHW (right) at 10 °C. For both compounds, the pH at which theionized state is favored is shown on the left and that for which theun-ionized state is favored is shown on the right. (C) Structure ofthe binding site, showing the direct interactions between NW and theGluA2 LBD [Protein Data Bank (PDB) entry 3RTW(32)]. (D) Structureof the binding site for NW showing the hydrogen bonding network associatedwith the nitrogen at position 3 of the willardiine ring in the chargedform (pH 6.5, PDB entry 3RTW(32)). On the left is thestructure and on the right a schematic. In the schematic, NW is coloredblue, the protein black, and water green and the H-bonds are coloredred. (E) Structure of NW bound to the GluA2 LBD obtained at pH 3.5(PDB entry 4Q30). At this pH, the ring is largely uncharged. The formatting of thispanel is similar to that of panel D. Note the change in the H-bondingnetwork. The potential H-bond between the hydroxyl of S654 and theuracil carbonyl is shown with a question mark because the distancebetween the two oxygens is relatively long, 3.5 Å, and the anglebetween this H-bond and the H-bond with the water is not favorable(80°).
Mentions: The influence ofsubstituent electronegativity on the thermodynamicparameters suggested that the differing pKa of the uracil ring, due to the different analogues in position 5,may affect the thermodynamics of binding. While the pKa of all the halogenated uracils is ∼8, those ofHW and NW are approximately 10 and 6, respectively.29,30 This means that willardiine and the halogenated analogues were,at physiological pH, in mostly uncharged (protonated) states (Figure 2A). NW, however, was in a mostly charged (deprotonated)state. The GluA2 LBD is well-behaved between pH 4 and 10, with minorchanges in the NMR 1H–15N HSQC spectrum.9 We tested the possibility that the charged stateof the ligand could affect the enthalpy versus entropy distributionby changing the pH. Thus, binding of largely uncharged (protonateduracil ring) NW was examined at pH 4, and binding of largely charged(deprotonated uracil ring) HW was examined at pH 10.

Bottom Line: The binding of the charged form is largely driven by enthalpy, while that of the uncharged form is largely driven by entropy.This is due at least in part to changes in the hydrogen bonding network within the binding site involving one water molecule.This work illustrates the importance of charge to the thermodynamics of binding of agonists and antagonists to AMPA receptors and provides clues for further drug discovery.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Medicine, Cornell University , Ithaca, New York 14853, United States.

ABSTRACT
Understanding the thermodynamics of binding of a lead compound to a receptor can provide valuable information for drug design. The binding of compounds, particularly partial agonists, to subtypes of the α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptor is, in some cases, driven by increases in entropy. Using a series of partial agonists based on the structure of the natural product, willardiine, we show that the charged state of the ligand determines the enthalpic contribution to binding. Willardiines have uracil rings with pKa values ranging from 5.5 to 10. The binding of the charged form is largely driven by enthalpy, while that of the uncharged form is largely driven by entropy. This is due at least in part to changes in the hydrogen bonding network within the binding site involving one water molecule. This work illustrates the importance of charge to the thermodynamics of binding of agonists and antagonists to AMPA receptors and provides clues for further drug discovery.

Show MeSH