Kinetic and Thermodynamic Evaluation of Kynurenic Acid Binding to GluR1270–300 Polypeptide by Surface Plasmon Resonance Experiments

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• 作者：Ádám Juhász ; Edit Csapó ; Ditta Ungor ; Gábor K. Tóth ; László Vécsei ; Imre Dékány
• 刊名：Journal of Physical Chemistry B
• 出版年：2016
• 出版时间：August 18, 2016
• 年：2016
• 卷：120
• 期：32
• 页码：7844-7850
• 全文大小：400K
• 年卷期：0
• ISSN：1520-5207

文摘

This work clearly demonstrates an evaluation process that is easily performed and is simply based on the fitting of temperature-dependent surface plasmon resonance (SPR) sensorgrams to provide detailed thermodynamic characterization of biologically relevant interactions. The reversible binding of kynurenic acid (KYNA) on human glutamate receptor (GluR1) polypeptide (GluR1_270–300)-modified gold surface has been studied at various temperatures under physiological conditions by two-dimensional SPR experiments. The registered sensorgrams were fitted by using different kinetic models without application of any commercial software. Assuming that the association of GluR1_270–300–KYNA complex is first order in both reactants, the association (k_a) and dissociation (k_d) constants as well as the equilibrium constants (K_A) and the Gibbs free-energy change (ΔG°) were given at 10, 20, 30, and 40 °C. Moreover, the enthalpy (ΔH° = −27.91 kJ mol^–1), entropy (ΔS° = −60.33 J mol^–1 K^–1), and heat capacity changes (ΔC_p = −1.28 kJ mol^–1 K^–1) of the model receptor–ligand system were also calculated using a spreadsheet program. Negative values of ΔG° and ΔH° indicate the exothermic formation of a stable GluR1_270–300–KYNA complex, because the |ΔH| > |TΔS| relation suggests an enthalpy-driven binding process. The negative ΔH° and ΔS° values strongly support the formation of a salt bridge between KYNA and the positively charged residues of the polypeptide (Arg, Lys) at pH 7.4, confirmed by molecular docking calculations as well.