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Related Concept Videos

Relative Strengths of Conjugate Acid-Base Pairs02:29

Relative Strengths of Conjugate Acid-Base Pairs

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Brønsted-Lowry acid-base chemistry is the transfer of protons; thus, logic suggests a relation between the relative strengths of conjugate acid-base pairs. The strength of an acid or base is quantified in its ionization constant, Ka or Kb, which represents the extent of the acid or base ionization reaction. For the conjugate acid-base pair HA / A−, the ionization equilibrium equations and ionization constant expressions are
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Acid/Base Strengths and Dissociation Constants03:02

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The relative strength of an acid or base is the extent to which it ionizes when dissolved in water. If the ionization reaction is essentially complete, the acid or base is termed strong; if relatively little ionization occurs, the acid or base is weak. There are many more weak acids and bases than strong ones. The most common strong acids and bases are listed below:
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The Equilibrium Binding Constant and Binding Strength02:18

The Equilibrium Binding Constant and Binding Strength

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The equilibrium binding constant (Kb) quantifies the strength of a protein-ligand interaction. Kb can be calculated as follows when the reaction is at equilibrium:
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Complexation Equilibria: Overview01:23

Complexation Equilibria: Overview

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Complexation reactions take place when dative or coordinate covalent bonds form between metal ions and ligands. The compounds formed in these reactions are called coordination compounds. The number of bonds formed between the metal ion and the ligands is called its coordination number. Generally, most metal ions in an aqueous solution are solvated by water molecules and thus exist as aqua complexes.
The equilibrium constant of the complexation reaction is represented as the formation constant...
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Acid and Bases: Ka, pKa, and Relative Strengths02:35

Acid and Bases: Ka, pKa, and Relative Strengths

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This lesson delves into a critical aspect of the relative strengths of acids and bases. The strength of an acid is evaluated by the acid dissociation into its conjugate base and a hydronium ion in water. The complete dissociation of a strong acid is confirmed with a very high concentration of hydronium ions. As a result, an incomplete dissociation process affirms a weak acid. Therefore, the equilibrium is in the forward direction for strong acids and backward for weak acids in these reactions.
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EDTA: Conditional Formation Constant01:09

EDTA: Conditional Formation Constant

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Each EDTA molecule has six binding sites: four carboxyl groups and two amino groups. The fully protonated form of EDTA is represented as H6Y2+. However, it can exist in different forms, H5Y+, H4Y, H3Y−, H2Y2−, and HY3−, depending on the pH of the solution. In very basic solutions with pH > 10.17, the fully deprotonated form, Y4−, is the predominant species that readily complexes with metal ions in a 1:1 ratio.
For the equilibrium reaction of the metal with the...
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Related Experiment Video

Updated: Jun 12, 2025

Titration ELISA as a Method to Determine the Dissociation Constant of Receptor Ligand Interaction
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Simple methods to determine the dissociation constant, Kd.

Donghun Lee1, Juwon Kim1, Gwangrog Lee1

  • 1Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 34141, South Korea.

Molecules and Cells
|September 18, 2024
PubMed
Summary

We present a novel single-molecule fluorescence method to determine dissociation constants (Kd), offering precise insights into protein-substrate binding affinities. This technique enhances understanding of molecular interactions crucial for drug development.

Keywords:
Dissociation constantElectrophoretic mobility shift assayEnzymeKineticsProteinSingle-molecule fluorescence resonance energy transfer

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Last Updated: Jun 12, 2025

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Area of Science:

  • Biochemistry
  • Pharmacology
  • Molecular Biology

Background:

  • The dissociation constant (Kd) is essential for quantifying binding affinities in biological systems.
  • Understanding these affinities is critical for drug development and biological mechanism elucidation.

Purpose of the Study:

  • To introduce a single-molecule fluorescence resonance energy transfer (smFRET) method for determining Kd.
  • To compare the smFRET method with the conventional electrophoretic mobility shift assay (EMSA) for Kd determination.
  • To provide insights into the thermodynamic interactions between proteins and substrates.

Main Methods:

  • Utilizing single-molecule fluorescence resonance energy transfer (smFRET) with fluorescence labeling.
  • Employing the electrophoretic mobility shift assay (EMSA) as a conventional method.
  • Analyzing binding and dissociation kinetics based on fluorescence signals and protein-DNA interactions.

Main Results:

  • The smFRET method accurately measures binding and dissociation kinetics.
  • Demonstrates the capability of smFRET in characterizing thermodynamic interactions.
  • Highlights the intrinsic nature of protein-DNA interactions.

Conclusions:

  • The smFRET method represents a significant advancement for determining Kd.
  • This technique offers enhanced accuracy and insights into molecular interactions.
  • Improves capabilities in molecular biology and pharmacological research.