A spin label study of egg white avidin.

Area of Science:

• Biochemistry
• Protein-ligand interactions
• Biophysical chemistry

Background:

• Avidin is a tetrameric protein known for its high affinity for biotin.
• Understanding the precise topography of biotin binding sites is crucial for various applications.

Purpose of the Study:

• To investigate the spatial arrangement and interactions of biotin analogs within avidin's binding sites.
• To determine the distance between adjacent binding sites on the avidin molecule.

Main Methods:

• Utilized four distinct spin-labeled biotin analogs.
• Employed fluorescence spectroscopy, optical absorption spectroscopy, and electron spin resonance (ESR) spectroscopy.
• Analyzed ESR spectral data, including hyperfine splitting and dipolar interactions, to infer distances and binding dynamics.

Main Results:

• Spin-labeled biotin analogs bind to the same sites as biotin.
• ESR dipole-dipole interactions revealed distances of approximately 15.7-16.7 Å between adjacent spin labels.
• Observed distinct binding and dissociation kinetics for different spin-labeled analogs, suggesting specific interactions and potential site redistribution.

Conclusions:

• The avidin binding site for biotin is likely a cleft within a hydrophobic depression.
• The spatial arrangement of binding sites allows for specific interactions and influences the binding dynamics of biotin analogs.
• Spin labeling provides a powerful tool for probing protein-ligand interactions and binding site topography.