Equilibrium Binding Model for CpG DNA-Dependent Dimerization of Toll-like Receptor 9 Ectodomain

Microbial
nucleic acids in the extracellular milieu are recognized
in vertebrates by Toll-like receptors (TLRs), one of the most important
families of innate immune receptors. TLR9 recognizes single-stranded
unmethylated CpG DNA in endosomes. DNA binding induces TLR9 dimerization
and activation of a potent inflammatory response. To provide insights
on how DNA ligands induce TLR9 dimerization, we developed a detailed
theoretical framework for equilibrium ligand binding, modeling the
binding of the ssDNA at the two main sites on the TLR9 ectodomain.
Light scattering and fluorescence anisotropy assays performed with
recombinant TLR9 ectodomain and a panel of agonistic and antagonistic
DNA ligands provide data that restrain the binding parameters, identify
the likely ligand binding intermediates, and suggest cooperative modes
of binding. This work brings us one step closer to establishing a
rigorous biochemical understanding of how TLRs are activated by their
ligands.