Doubling Forster Resonance Energy Transfer Efficiency in Proteins with Extrinsic Thioamide Probes: Implications for Thiomodified Nucleobases

Designing a potential protein-ligand pair is pivotal, not only to track the protein structure dynamics, but also to assist in an atomistic understanding of drug delivery. Herein, the potential of a small model thioamide probe being used to study albumin proteins is reported. By monitoring the Forster resonance energy transfer (FRET) dynamics with the help of fluorescence spectroscopic techniques, a twofold enhancement in the FRET efficiency of 2-thiopyridone (2TPY), relative to that of its amide analogue, is observed. Molecular dynamics simulations depict the relative position of the free energy minimum to be quite stable in the case of 2TPY through noncovalent interactions with sulfur, which help to enhance the FRET efficiency. Finally, its application is shown by pairing thiouracils with protein. It is found that the site-selective sulfur atom substitution approach and noncovalent interactions with sulfur can substantially enhance the FRET efficiency, which could be a potential avenue to explore in the design of FRET probes to study the structure and dynamics of biomolecules.

Automated summary

This study highlights the importance of designing protein-ligand pairs for tracking protein dynamics and assisting in drug delivery. Using a small model thioamide probe, the enhanced FRET efficiency in studying albumin proteins is demonstrated. Selective sulfur atom substitution and noncovalent interactions with sulfur are shown to substantially enhance FRET efficiency, offering a potential avenue for designing FRET probes for studying biomolecular structure and dynamics.