Molecular Modeling-Based Evaluation of hTLR10 and Identification of Potential Ligands in Toll-Like Receptor Signaling

Abstract

Toll-like receptors (TLRs) are pattern recognition receptors that recognize pathogens based on distinct molecular signatures. The human (h)TLR1, 2, 6 and 10 belong to the hTLR1 subfamilies, which are localized in the extracellular regions and activated in response to diverse ligand molecules. Due to the unavailability of the hTLR10 crystal structure, the understanding of its homo and heterodimerization with hTLR2 and hTLR1 and the ligand responsible for its activation is limited. To improve our understanding of the TLR10 receptor-ligand interaction, we used homology modeling to construct a three dimensional (3D) structure of hTLR10 and refined the model through molecular dynamics (MD) simulations. We utilized the optimized structures for the molecular docking in order to identify the potential site of interactions between the homo and heterodimer (hTLR10/2 and hTLR10/1). The docked complexes were then used for interaction with ligands (Pam3CSK4 and PamCysPamSK4) using MOE-Dock and ASEDock. Our docking studies have shown the binding orientations of hTLR10 heterodimer to be similar with other TLR2 family members. However, the binding orientation of hTLR10 homodimer is different from the heterodimer due to the presence of negative charged surfaces at the LRR11-14, thereby providing a specific cavity for ligand binding. Moreover, the multiple protein-ligand docking approach revealed that Pam3CSK4 might be the ligand for the hTLR10/2 complex and PamCysPamSK4, a di-acylated peptide, might activate hTLR10/1 hetero and hTLR10 homodimer. Therefore, the current modeled complexes can be a useful tool for further experimental studies on TLR biology.

IgM+IgD+CD27+ B cells are markedly reduced in IRAK-4–, MyD88-, and TIRAP- but not UNC-93B–deficient patients

“…Along this line, we addressed whether the 2 CD27 ϩ subsets have different effector profiles in response to various TLR agonists, including PamCys-(Pam)SK4, a proposed TLR10-specific ligand based on modeling studies. 33 Unfortunately, no response either in terms of cytokine/ chemokine secretion or cellular proliferation was observed for this agonist, thus precluding to get insight as to whether TLR10 may have a unique function for IgM ϩ IgD ϩ CD27 ϩ cells. We found that TLR2/6, TLR7, and TLR9 triggering activated all splenic subsets to produce cytokines, chemokines, and hematopoietic growth factors.…”

“…Sorted B cells were incubated in 96-well flat-bottom plates (BD Biosciences) at a concentration of 2 ϫ 10 5 cells/200L in RPMI1640/Glutamax/ PenStrep (Gibco-Invitrogen) and 10% FCS (FetalClone I; HyClone) and were stimulated with different TLR agonists, used at the following final concentrations: CpG ODN 2006 (TLR9 agonist, Sigma-Aldrich) at 6 g/ mL; flagellin (TLR5 agonist) at 0.5 g/mL; resiquimod or R848 (TLR7 and 8 agonist), Pam2CSK4 (TLR2/6 agonist), and Pam3CSK4 (TLR1/2 agonist; all from Invivogen) at 1 g/mL; PamCys(Pam)-SK4 (a putative TLR10 agonist, 33 EMC Microcollections) at 1 g/mL; and poly-IC (polyinosine-polycytidic acid, a TLR3 agonist, GE Healthcare) at 25 ng/L. The quantitative determination of secreted cytokines/chemokines was performed on 96-well supernatants collected after 4 days in culture via the use of multiplex cytokine assays (Human Cytokine 25-plex panel from Invitrogen or Bio-Plex Pro Human Cytokine 27-Plex Panel from Bio-Rad) read on a Bio-Plex 200 system (Bio-Rad).…”

“…Two attempts at inducing TLR10 signaling were made: first, by the use of PamCys(Pam)-SK4, which was proposed as a potential specific ligand of TLR10 on the basis of modeling studies and might activate human TLR10/1 hetero-and TLR10 homodimers, 33 and second, through TLR10 cross-linking via the use of anti-TLR10 antibodies. However, both approaches were unable to trigger cytokine/chemokine secretion (or cell proliferation) from CD27 Ϫ or CD27 ϩ B-cell subsets.…”

Dendritic Cells Interactions with the Immune System – Implications for Vaccine Development

“…There is low evidence about the ligands binding TLR10. A study carried out by Govindaraj et al [62] showed that PamCysPamSK4, a di-acylated peptide, might activate the human TLR10/1 hetero and TLR10 homodimer, while Pam3CSK4 might be recognized by the TLR10/2 heterodimer.…”

Structure and dynamic behavior of Toll‐like receptor 2 subfamily triggered by malarial glycosylphosphatidylinositols of Plasmodium falciparum

“…Homology models of TLR10 complexes were built and refined through molecular dynamics (MD) simulations to predict the protein–ligand complexes hTLR2‐hTLR10‐Pam 3 CSK 4 , hTLR1‐hTLR10‐PamCysPamSK 4 and hTLR10‐hTLR10‐PamCysPamSK 4 28 . X‐ray crystallography studies of dimeric complexes of TLR2 in combination with TLR1, which recognizes Pam 3 CSK 4 , and another combination of TLR2 with TLR6, which recognizes Pam 2 CSK 4 , have dissected ECDs of the TLR2 subfamily 17.…”