Modelling and validation of VIP neuropeptide binding to the CRTH2

Eosinophilic esophagitis (EoE) is a chronic, immune-mediated inflammatory condition of the esophagus, characterized by eosinophilic infiltration driven by a T-helper type-2 (Th2) immune response. Key cytokines (IL-4, IL-5, IL-13) facilitate eosinophil recruitment, survival and activation, while prostaglandin D2 (PGD₂) signaling via CRTH2 promotes eosinophil chemotaxis. Experimental studies suggest a functional association between Vasoactive Intestinal Peptide (VIP) and CRTH2, where VIP may upregulate CRTH2 expression and enhance eosinophil migration. However, the precise molecular basis of this interaction, particularly the physical binding interface, remains unclear. This thesis aims to explore the structural basis of the VIP-CRTH2 interaction using advanced in silico approaches. Structural characterization of full-length and N-terminal truncated CRTH2 variants via CAVER 3.0 revealed the N-terminal lid as a critical steric gate, suggesting partial VIP unfolding for entry. Molecular docking simulations of VIP-CRTH2 complexes using PyRosetta consistently indicated shallow, peripheral VIP-CRTH2 interactions, primarily at the extracellular surface, without deep pocket engagement. AlphaFold3-based structure prediction models with subsequent contact analysis via RING 4.0 showed consistent interaction signals, with the polar sub-pocket being interaction-dense due to hydrogen bonds and ionic interactions. However, moderate interface confidence (ipTM < 0.6) and a prevalence of van der waals over strong polar interactions, alongside AlphaFold3's N-terminal VIP preference, suggest limitations in capturing specific polar interactions and raise concerns about specificity. This work establishes a structural foundation and insights for future Molecular Dynamic studies and alternative docking simulations with CABS-dock of the VIP-CRTH2 interaction.