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Cooperativity in Binding Processes: New Insights from Phenomenological Modeling.

Cattoni DI, Chara O, Kaufman SB, González Flecha FL - PLoS ONE (2015)

Bottom Line: Here we analyze the simplest phenomenological model that can account for cooperativity (i.e. ligand binding to a macromolecule with two binding sites) by generating equilibrium binding isotherms from deterministically simulated binding time courses.We show that the Hill coefficients determined for cooperative binding, provide a good measure of the Gibbs free energy of interaction among binding sites, and that their values are independent of the free energy of association for empty sites.We also conclude that although negative cooperativity and different classes of binding sites cannot be distinguished at equilibrium, they can be kinetically differentiated.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de Biofísica Molecular, Instituto de Química y Fisicoquímica Biológicas, Universidad de Buenos Aires - CONICET, Buenos Aires, Argentina.

ABSTRACT
Cooperative binding is one of the most interesting and not fully understood phenomena involved in control and regulation of biological processes. Here we analyze the simplest phenomenological model that can account for cooperativity (i.e. ligand binding to a macromolecule with two binding sites) by generating equilibrium binding isotherms from deterministically simulated binding time courses. We show that the Hill coefficients determined for cooperative binding, provide a good measure of the Gibbs free energy of interaction among binding sites, and that their values are independent of the free energy of association for empty sites. We also conclude that although negative cooperativity and different classes of binding sites cannot be distinguished at equilibrium, they can be kinetically differentiated. This feature highlights the usefulness of pre-equilibrium time-resolved strategies to explore binding models as a key complement of equilibrium experiments. Furthermore, our analysis shows that under conditions of strong negative cooperativity, the existence of some binding sites can be overlooked, and experiments at very high ligand concentrations can be a valuable tool to unmask such sites.

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Comparison between negative cooperativity and different sites in pre-equilibrium conditions.(A) Time course of site occupation for a macromolecule with two identical sites for negative cooperativity (continuous lines) and two classes of binding sites without interactions (dash-dotted lines) for the following initial ligand concentrations (μM): 300 (red), 100 (orange), 50 (green), 25 (turquoise), 10 (blue) and 5 (violet). Time courses simulations were obtained under the same conditions as described in Fig 5. (B) The difference Δ〈n〉 = 〈n〉coop—〈n〉diff sites was calculated for each ligand concentration and represented as a function of time. For clarity reasons only 6 representative time courses traces that gave rise to the equilibrium data points of Fig 5 are shown.
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pone.0146043.g006: Comparison between negative cooperativity and different sites in pre-equilibrium conditions.(A) Time course of site occupation for a macromolecule with two identical sites for negative cooperativity (continuous lines) and two classes of binding sites without interactions (dash-dotted lines) for the following initial ligand concentrations (μM): 300 (red), 100 (orange), 50 (green), 25 (turquoise), 10 (blue) and 5 (violet). Time courses simulations were obtained under the same conditions as described in Fig 5. (B) The difference Δ〈n〉 = 〈n〉coop—〈n〉diff sites was calculated for each ligand concentration and represented as a function of time. For clarity reasons only 6 representative time courses traces that gave rise to the equilibrium data points of Fig 5 are shown.

Mentions: When comparing the time courses of ligand binding significant differences between both sets of simulated data can be observed (Fig 6A) allowing to identify the corresponding binding model. To highlight this, the difference between the average numbers of occupied sites (Δ〈n〉 = 〈n〉coop−〈n〉diff. sites) for each total ligand concentration is depicted in Fig 6B. It can be observed that differences, initially small, reach a maximal value at different times (depending on ligand concentration) and become indistinguishable when approaching to equilibrium (Fig 6A and 6B).


Cooperativity in Binding Processes: New Insights from Phenomenological Modeling.

Cattoni DI, Chara O, Kaufman SB, González Flecha FL - PLoS ONE (2015)

Comparison between negative cooperativity and different sites in pre-equilibrium conditions.(A) Time course of site occupation for a macromolecule with two identical sites for negative cooperativity (continuous lines) and two classes of binding sites without interactions (dash-dotted lines) for the following initial ligand concentrations (μM): 300 (red), 100 (orange), 50 (green), 25 (turquoise), 10 (blue) and 5 (violet). Time courses simulations were obtained under the same conditions as described in Fig 5. (B) The difference Δ〈n〉 = 〈n〉coop—〈n〉diff sites was calculated for each ligand concentration and represented as a function of time. For clarity reasons only 6 representative time courses traces that gave rise to the equilibrium data points of Fig 5 are shown.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4696654&req=5

pone.0146043.g006: Comparison between negative cooperativity and different sites in pre-equilibrium conditions.(A) Time course of site occupation for a macromolecule with two identical sites for negative cooperativity (continuous lines) and two classes of binding sites without interactions (dash-dotted lines) for the following initial ligand concentrations (μM): 300 (red), 100 (orange), 50 (green), 25 (turquoise), 10 (blue) and 5 (violet). Time courses simulations were obtained under the same conditions as described in Fig 5. (B) The difference Δ〈n〉 = 〈n〉coop—〈n〉diff sites was calculated for each ligand concentration and represented as a function of time. For clarity reasons only 6 representative time courses traces that gave rise to the equilibrium data points of Fig 5 are shown.
Mentions: When comparing the time courses of ligand binding significant differences between both sets of simulated data can be observed (Fig 6A) allowing to identify the corresponding binding model. To highlight this, the difference between the average numbers of occupied sites (Δ〈n〉 = 〈n〉coop−〈n〉diff. sites) for each total ligand concentration is depicted in Fig 6B. It can be observed that differences, initially small, reach a maximal value at different times (depending on ligand concentration) and become indistinguishable when approaching to equilibrium (Fig 6A and 6B).

Bottom Line: Here we analyze the simplest phenomenological model that can account for cooperativity (i.e. ligand binding to a macromolecule with two binding sites) by generating equilibrium binding isotherms from deterministically simulated binding time courses.We show that the Hill coefficients determined for cooperative binding, provide a good measure of the Gibbs free energy of interaction among binding sites, and that their values are independent of the free energy of association for empty sites.We also conclude that although negative cooperativity and different classes of binding sites cannot be distinguished at equilibrium, they can be kinetically differentiated.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de Biofísica Molecular, Instituto de Química y Fisicoquímica Biológicas, Universidad de Buenos Aires - CONICET, Buenos Aires, Argentina.

ABSTRACT
Cooperative binding is one of the most interesting and not fully understood phenomena involved in control and regulation of biological processes. Here we analyze the simplest phenomenological model that can account for cooperativity (i.e. ligand binding to a macromolecule with two binding sites) by generating equilibrium binding isotherms from deterministically simulated binding time courses. We show that the Hill coefficients determined for cooperative binding, provide a good measure of the Gibbs free energy of interaction among binding sites, and that their values are independent of the free energy of association for empty sites. We also conclude that although negative cooperativity and different classes of binding sites cannot be distinguished at equilibrium, they can be kinetically differentiated. This feature highlights the usefulness of pre-equilibrium time-resolved strategies to explore binding models as a key complement of equilibrium experiments. Furthermore, our analysis shows that under conditions of strong negative cooperativity, the existence of some binding sites can be overlooked, and experiments at very high ligand concentrations can be a valuable tool to unmask such sites.

Show MeSH
Related in: MedlinePlus