Name: how equilibrium binding constant affects binding
Uploaded: 2023-04-30T18:28:41.000Z
Duration: 2 min 18 s
Channel: JoVE

Dans la cellule, les protéines entrent en collision aléatoire avec d'autres molécules, comme d'autres protéines, des acides nucléiques et des petites molécules ligands. Si la liaison est non-spécifique, peu d'interactions non-covalentes entre les molécules entraînent une brève association. Si un ligand spécifique se lie à la protéine, il forme de vastes interactions non-covalentes le long des surfaces complémentaires.Ces complexes sont stables, restant longtemps liés avant de se dissocier. La force d'une interaction de liaison est rapportée en termes de sa constante d'équilibre, Kb, également appelée liaison ou constante d'association. Le Kb peut être calculé à partir du rapport de la concentration du complexe protéine-ligand sur les concentrations de protéines non-liées et de ligands trouvés à l'équilibre.Étant donné sa relation avec le changement d'énergie libre dû à la liaison, une valeur impotante de Kb signifie une forte diminution du delta G indiquant une forte affinité entre la protéine et le ligand. Deux processus concurrents sont importants pour la liaison protéine-ligand. L'association d'une protéine et d'un ligand pour former un complexe et la dissociation du complexe en réactifs.La constante d'association K on est une mesure du nombre d'événements de liaison par seconde entre une protéine et son ligand. Il peut être utilisé pour calculer le taux de liaison du ligand à la protéine à une concentration donnée. Inversement, K off est une mesure du nombre d'événements de dissociation.Il peut être utilisé pour calculer la vitesse à laquel un complexe se défait lorsque le taux d'association est égal au taux de dissociation et que l'équilibre est atteint, où les concentrations de produits et de réactifs restent constantes. Ainsi, à l'équilibre, le K on multiplié par le produit des concentrations d'équilibre de la protéine et du ligand, est égal au K off multiplié par la concentration d'équilibre du complexe protéine-ligand. La réorganisation de cette expression montre également que le rapport de K on sur K off est égal au Kb à l'équilibre.

<ol>
	<li>Pollard, Thomas D. « Un guide pour des tests de liaison simples et informatifs. » Biologie moléculaire de la cellule 21, n° 23 (2010) : 4061-4067.</li>
	<li>Hulme, Edward C., et Mike A. Trevethick. « Tests de liaison de ligand à l’équilibre : validation et interprétation. » British journal of pharmacology 161, n° 6 (2010) : 1219-1237.</li>
	<li>Medina-Franco, José L., Oscar Méndez-Lucio et Karina Martinez-Mayorga. « L’interaction entre la modélisation moléculaire et la chimio-informatique pour caractériser les paysages d’interactions protéine-ligand et protéine-protéine pour la découverte de médicaments. » Dans Advances in protein chemistry and structural biology, vol. 96, pp. 1-37. Presse académique, 2014.</li>
</ol>

la constante d'équilibre des liaisons et la force des liaisons

The Equilibrium Binding Constant and Binding Strength

La constante de liaison à l'équilibre (Kb) quantifie la force d'une interaction protéine-ligand. La constante de liaison à l'équilibre peut être calculé comme suit lorsque la réaction est à l'équilibre :

<img alt="Eq1" src="/files/ftp_upload/11513/Equation 1.png" style="text-align: center; width: 650px;" />

où P et L sont respectivement la protéine et le ligand non liés, et PL est le complexe protéine-ligand.

Comme la quantité de ligand lié est également liée au taux de liaison du ligand, les expériences peuvent également déterminer Kb en examinant les taux d'association protéine-ligand (kon) et de dissociation (koff) en utilisant le rapport suivant :

<img alt="Eq1" src="/files/ftp_upload/11513/Equation 2.png" style="text-align: center; width: 650px;" />

Ainsi, deux catégories d’analyses de liaison sont utilisées pour déterminer la constante de liaison à l’équilibre : celles qui mesurent les concentrations d’équilibre et celles qui mesurent la cinétique d’une réaction. Dans ce cas, la réaction doit être à l’équilibre au moment de la mesure.

La méthode de détermination des concentrations à l'équilibre dépend de la sensibilité souhaitée et de la facilité de détection du signal. Pour ces raisons, les analyses spectroscopiques sont les plus largement utilisées. Dans ces expériences, la réaction produit un changement d'absorbance d'un réactif ou d'un produit à une longueur d'onde donnée, détectable par un spectrophotomètre UV-Vis. Alternativement, le réactif ou le produit peut être marqué avec une sonde fluorescente ou peut contenir un fluorophore intrinsèque. Ensuite, la progression de la réaction peut être mesurée à partir du changement de fluorescence. Ces tests sont effectués en faisant varier les concentrations d’un réactif tandis que le reste de l’expérience reste constant. Les résultats peuvent ensuite être représentés graphiquement et analysés avec diverses méthodes d'ajustement de courbe.

Les interactions entre protéines et ligands sont également étudiées à l’aide de diverses techniques biochimiques et spectroscopiques. L'analyse structurale, utilisant la cristallographie aux rayons X et la spectroscopie RMN, aide à prédire les interactions protéine-ligand grâce à des simulations moléculaires. Des approches théoriques et informatiques, telles que les études d'amarrage protéine-ligand, sont largement utilisées pour caractériser la position et les interactions de ligands de petites molécules, y compris les candidats médicaments. La conception de médicaments assistée par ordinateur constitue une alternative rapide et peu coûteuse pour accélérer le rythme des tests conventionnels de médicaments par essais et erreurs.

La constante d'équilibre des liaisons et la force des liaisons

La constante de liaison à l’équilibre (Kb) quantifie la force d’une interaction protéine-ligand. Kb peut être calculé comme suit lorsque la réaction est à ...

the-equilibrium-binding-constant-and-binding-strength

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how equilibrium binding constant affects binding

The equilibrium binding constant describes how strongly two molecules bind at equilibrium. It is a useful way to compare binding strength in molecular interactions.

This topic focuses on how binding is measured and what the equilibrium binding constant means. A higher value indicates stronger binding, while a lower value indicates weaker binding. Understanding this constant helps explain how tightly molecules stay together under equilibrium conditions.

The page also connects the binding constant to real binding strength. It shows why this value matters when comparing different molecular pairs. For high school students, it provides a clear link between a chemistry concept and molecular behavior.

How Equilibrium Binding Constant Affects Binding

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the equilibrium binding constant and binding strength

The equilibrium binding constant describes how strongly two molecules bind at equilibrium. It is a useful way to compare binding strength in molecular interactions. This topic focuses on how binding is measured and what the equilibrium binding constant means. A higher value indicates stronger binding, while a lower value indicates weaker binding. Understanding this constant helps explain how tightly molecules stay together under equilibrium conditions. The page also connects the binding...

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key terms and concepts

Learn how the equilibrium binding constant reflects binding strength and why higher or lower values matter in molecular interactions at equilibrium.

How Equilibrium Binding Constant Affects Binding | JoVE K12

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How Equilibrium Binding Constant Affects Binding

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