Hydrogen bonds play crucial roles in functional biomolecular dynamics. It is suggested that non-conventional hydrogen bonds engaging π electrons are prevalent in proteins. The experimental support for their existence is however limited. Here, we provide direct NMR spectroscopic evidence for the existence of an NH-π interaction in an intrinsically disordered peptide (E22G-Aβ40). In particular, we demonstrate the correlation between the amide proton of a glycine residue (Gly22) and the aromatic carbons of its preceding Phe20 through π hydrogen bond-mediated scalar coupling between them, as predicted by density functional theory calculations. Our results present a proof-of-principle example of NH-π interactions in an intrinsically disordered protein (IDP) and suggest the potential prevalence of π hydrogen bonds on the surface of IDPs. Direct experimental verification of NH-π interactions in folded proteins remains for future studies.
© 2025. The Author(s).