Effects of thermodynamic nonideality in ligand binding studies

Area of Science:

• Biophysical Chemistry
• Macromolecular Science
• Thermodynamics

Background:

• Accurate characterization of small ligand-macromolecule interactions is crucial in biochemistry.
• Thermodynamic nonideality can influence binding measurements, necessitating careful consideration.

Purpose of the Study:

• To quantitatively assess the impact of thermodynamic nonideality on ligand-macromolecule binding.
• To formulate expressions for binding functions accounting for nonideality effects.
• To evaluate the applicability of standard binding equations under nonideal conditions.

Main Methods:

• Equilibrium dialysis to determine ligand concentration in dialysis equilibrium.
• Ultrafiltration and rate of dialysis methods to measure unbound ligand concentration.
• Formulation of binding functions incorporating molar volume, charge-charge interaction, and covolume contributions.
• Experimental studies using tryptophan and methyl orange with bovine serum albumin.

Main Results:

• Explicit expressions for binding functions were derived, considering composition-dependent nonideality.
• Experimental studies explored the magnitude of these nonideality effects.
• For equilibrium dialysis and frontal gel chromatography, acceptor-concentration dependence due to nonideality was found to be negligible after Donnan correction.
• For ultrafiltration and rate of dialysis, the assumption of unity for activity coefficient ratios is generally valid.

Conclusions:

• Standard binding equations are appropriate for analyzing results from equilibrium dialysis and frontal gel chromatography, even with nonideality.
• Nonideality effects on binding functions are unlikely to be significant, providing reassurance for common experimental methods.
• The study simplifies the interpretation of ligand-macromolecule binding data by confirming the general applicability of ideal system models.