Structural and energetic requirements for a second binding site at the dimeric β-lactoglobulin interface.

Area of Science:

• Protein structure and dynamics
• Molecular biophysics
• Biochemistry

Background:

• Bovine beta-lactoglobulin (βlg) primarily binds hydrophobic molecules in its calyx.
• A secondary binding site for vitamin D3 (VD3) exists at the βlg dimeric interface, even in denatured states.
• Previous studies suggest structural requirements for VD3 binding to monomeric and dimeric βlg.

Purpose of the Study:

• To investigate the dynamic interaction of VD3 with monomeric and dimeric βlg.
• To determine the energetic differences of VD3 binding at the calyx versus the dimeric interface.
• To elucidate the role of βlg structure in VD3 complex stability.

Main Methods:

• Blind docking simulations
• 2-microsecond molecular dynamics simulations
• Binding free energy calculations (MM/GBSA)

Main Results:

• The dimeric structure of βlg is essential for stable VD3 complex formation at the dimeric interface.
• Monomeric βlg can bind VD3 at the dimeric interface, but with marginally favorable binding free energy due to entropic contributions.
• Per-residue decomposition analysis identified key residues stabilizing the VD3-βlg complexes.

Conclusions:

• The dimeric interface is a significant secondary binding site for VD3 in βlg.
• VD3 binding influences the dimerization process of βlg.
• Understanding these interactions provides insights into protein-ligand dynamics and stability.