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

In the cell, proteins randomly collide with other molecules, such as other proteins, nucleic acids, and small-molecule ligands.
If the binding is non-specific, few non-covalent interactions between the molecules result in a brief association.
If a specific ligand binds to the protein, it forms extensive non-covalent interactions along complementary surfaces. Such complexes are stable, staying bound for a long time, before dissociating.
The strength of a binding interaction is reported in terms of its equilibrium constant, Kb, also called the binding or association constant. 
Kb can be calculated from the ratio of the concentration of protein-ligand complex over the concentrations of unbound protein and ligand found at equilibrium. 
Given its relation with the free energy change due to binding, a large Kb means a large decrease in delta G, indicating strong affinity between protein and ligand.
Two competing processes are important for protein-ligand binding: the association of a protein and a ligand to form a complex and the dissociation of the complex into the reactants.
The association constant, kon, is a measure of the number of binding events per second between a protein and its ligand; it can be used to calculate the rate of ligand binding to the protein at a given concentration. 
Conversely, koff, is a measure of the number of dissociation events; it can be used to calculate how quickly the complex comes apart.
When the rate of association equals the rate of dissociation, an equilibrium is reached, where the net concentrations of products and reactants remain constant.
Thus, at equilibrium, kon times the product of the equilibrium concentrations of the protein and the ligand equals koff times the equilibrium concentration of the protein-ligand complex.
Rearranging this expression also shows that the ratio of kon to koff equals Kb at equilibrium.

the equilibrium binding constant and binding strength

The Equilibrium Binding Constant and Binding Strength

De evenwichtsbindingsconstante (K_bKb) kwantificeert de sterkte van een eiwit-ligand-interactie. K_bligandinteractie. Kb kan als volgt worden berekend alswanneer de reactie in evenwicht is, volgens de vergelijking:

<img alt="Eq1" src="/files/ftp_upload/11513/Equation 1.png" style="text-align: center; max-width: 100%;">

waarbij P en L respectievelijk het ongebonden eiwit en ligand zijnvoorstellen, en PL het eiwit-ligandcomplex is.

Omdat de hoeveelheid gebonden ligand ook gerelateerd is aan de snelheid van ligandbinding, kunnen experimenten ook K_b bepalenkan Kb experimenteel worden bepaald door de snelheid van eiwit-ligand-associatie (k_onligandassociatie (kon) en dissociatie (k_offkoff) te onderzoeken met behulp van de volgende meten volgens de verhouding:

<img alt="Eq1" src="/files/ftp_upload/11513/Equation 2.png" style="text-align: center; max-width: 100%;">

Er worden duszijn twee categorieën bindingstesten gebruiktbindingsassays om de evenwichtsbindingsconstante te bepalen: degenemethoden die evenwichtsconcentraties meten en degenemethoden die de kinetiek van eende reactie meten. In dat gevalanalyseren. Bij de eerste moet de reactie op het moment van de meting in evenwicht zijn.

De methode voor het bepalen van de om evenwichtsconcentraties te bepalen hangt af van de gewenste gevoeligheid en het gemakde eenvoud van signaaldetectie. Om deze redenen worden spectroscopische testenassays het meest gebruikt. In deze experimenten produceertveroorzaakt de reactie een absorptieverandering van een reactantreagens of een product bij een bepaaldespecifieke golflengte, detecteerbaardie wordt gedetecteerd met een UV-Vis-spectrofotometer. Als alternatiefAlternatief kan de reactanteen reagens of het product worden gelabeldgetagd met een fluorescerendefluorescente probe of kan deze een intrinsieke fluorofoorfluorofore groep bevatten. VervolgensDe voortgang van de reactie kan het reactieverloopdan worden gemeten aan de hand vandoor de verandering in fluorescentie. -intensiteit.

Deze testsassays worden uitgevoerd door de concentratiesconcentratie van één reactantreagens te variëren, terwijl de rest van het experimentoverige experimentele omstandigheden constant wordt gehouden.blijven. De verkregen resultaten kunnen vervolgens in een grafiek worden weergegeven en worden geanalyseerd met verschillende curve-fittingmethodenfitting methoden.

Interacties tussen eiwitten en liganden worden ook bestudeerd met behulp van een verscheidenheid aaneen reeks biochemische en spectroscopische technieken. Structurele analyse, met behulp vanzoals röntgenkristallografie en NMR-spectroscopie, helpt bij het voorspellen van eiwit-ligand-interacties door middel vanligandinteracties via moleculaire simulaties. Theoretische en computationele benaderingen, zoals onderzoeken naar de koppeling van eiwit-ligandenligand dockingstudies, worden veelvuldigveel gebruikt om de positie en interacties van liganden van kleine moleculen, moleculaire liganden, inclusief kandidaat-geneesmiddelenmedicijnkandidaten, te karakteriseren. Computerondersteund geneesmiddelenontwerp isComputergestuurde medicijnontwikkeling biedt een snelsnelle en goedkoop alternatiefkosteneffectieve manier om het tempoproces van conventionele proefondervindelijke geneesmiddelentestsmedicijnscreening te versnellen.

The equilibrium binding constant (Kb) quantifies the strength of a protein-ligand interaction. Kb can be calculated as follows when the reaction is at ...

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

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De evenwichtsbindingsconstante en bindingssterkte

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 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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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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