Palomino-Hernandez, O. | bioRxiv Science

bioRxiv2025

A hydrophobic cluster controls long-range allostery in the TRMT2A RNA recognition motif

TRMT2A has emerged as a disease-modifying target in polyglutamine (PolyQ) models, yet the conformational preferences and allostery of its RNA recognition motif (RRM) remain poorly resolved. Here we combine extensive atomistic molecular dynamics with Markov state modeling (BHMSM), transition path theory, and structure-based pocket analysis to map the conformational landscape of the human TRMT2A RRM. We resolve six metastable states and show that a hydrophobic cluster centered on F92-W134-L133 governs their interconversion. We further identify residues contributing to RNA strand recognition and reveal state-specific cryptic pockets consistent with reported TRMT2A RRM small molecule inhibitors. Together, these results support a hinge-gate model in which a soft, defectenabled 2 segment and a loop 5 hydrophobic cluster coordinate long-range communication between the RNP face and opposite side, yielding testable mutational predictions and state-specific opportunities for allosteric control of TRMT2A in polyQ disease contexts.

biophysics↗

bioRxiv2024

Protonation effects in protein-ligand complexes - a case study of endothiapepsin and pepstatin A with computational and experimental methods

1Protonation states serve as an essential molecular recognition motif for biological processes. Their correct consideration is key to successful drug design campaigns, since chemoinformatic tools usually deal with default protonation states of ligands and proteins and miss atypical protonation states. The protonation pattern for the Endothiapepsin/PepstatinA (EP/pepA) complex is investigated using different dry lab and wet lab techniques. ITC experiments revealed an uptake of more than one mole of protons upon pepA binding to EP. Since these experiments were performed at physiological conditions (and not at pH=4 at which a large variety of crystal structures is available), a novel crystal structure at pH=7.6 was determined. This crystal structure showed that only modest structural changes occur upon increasing the pH value. This lead to computational studies to reveal the exact location of the protonation event. Both computational studies could reveal a significant pKa shift resulting in non-default protonation state and that the catalytic dyad is responsible for the uptake of protons. This study shows that assessing protonation states for two separate systems (protein and ligand) might result in the incorrect assignment of protonation states and hence incorrect calculation of binding energy.

molecular biology↗