Tryptophan as a molecular microswitch: Integrating BioLiP statistical insights with MD simulations of 5-HT₂ receptors

Abstract

Tryptophan residues play a critical role in protein-ligand recognition owing to their unique aromatic character, high polarizability, and ability to participate in diverse noncovalent interactions, including π-π stacking, cation-π interactions, hydrogen bonding, and hydrophobic contacts. In this study, a two-stage approach was employed to elucidate the functional significance of tryptophan residues within ligand-binding sites across diverse protein systems. First, a large-scale statistical analysis was performed using a curated dataset of protein-ligand complexes extracted from the BioLiP database. The dataset was analysed with respect to enzymatic classification and Gene Ontology enrichment, revealing a pronounced enrichment of tryptophan residues in the binding sites of hydrolases, oxidoreductases, and transferases, as well as strong associations with metal-ion binding, redox-related functions, and membrane or cytoplasmic localization. Within the G protein-coupled receptor (GPCR) superfamily, rhodopsin-like class A receptors were identified as the most prominently represented group featuring tryptophan-mediated ligand interactions. In the second stage, molecular dynamics simulations were conducted for selected serotonin 5-HT₂ receptor subtypes (5-HT₂A, 5-HT₂B, and 5-HT₂C) to provide atomistic insights into the role of the conserved W6.48 residue. The simulations reveal a stable spatial relationship between the ligand and W6.48, characterized by distances consistent with aromatic anchoring, supporting its role in maintaining ligand orientation and binding pocket organization. Collectively, these results highlight tryptophan as a key determinant of ligand recognition and stabilization across both enzymatic and receptor-mediated systems.