Probing subtle conformational changes induced by phosphorylation and point mutations in the TIR domains of TLR2 and TLR3.

Extensive research performed on Toll-like receptor (TLR) signaling has identified residues in the Toll/interleukin-1 receptor (TIR) domains that are essential for its proper functioning. Among these residues, those in BB loop are particularly significant as single amino acid mutations in this region can cause drastic changes in downstream signaling. However, while the effect of these mutations on the function is well studied (like the P681H mutation in TLR2, the A795P mutation in TLR3, and the P714H mutation in TLR4), their influence on the dynamics and inter-residue networks is not well understood. The effects of local perturbations induced by these mutations could propagate throughout the TIR domain, influencing interactions with other TIR domain-containing proteins. The identification of these subtle changes in inter-residue interactions can provide new insights and structural rationale for how single-point mutations cause drastic changes in TIR-TIR interactions. We employed molecular dynamics simulations and protein structure network (PSN) analyses to investigate the structural transitions with special emphasis on TLR2 and TLR3. Our results reveal that phosphorylation of the Tyr 759 residue in the TIR domain of TLR3 introduces rigidity to its BB loop. Subtle differences in the intra BB loop hydrogen bonding network between TLR3 and TLR2 are also observed. The PSN analyses indicate that the TIR domain is highly connected and pinpoints key differences in the inter-residue interactions between the wild-type and mutant TIR domains, suggesting that TIR domain structure is prone to allosteric effects, consistent with the current view of the influence of allostery on TLR signaling.