Investigating the binding of fluorescent probes to a trypanosomal-tRNA synthetase: A fluorescence spectroscopic and...

Given the high prevalence of Chagas disease in the Americas, we targeted the unique arginyl-tRNA synthetase of its causative agent Trypanosoma cruzi. Among their many possible uses, naphthalene-derived fluorescent ligands, such as ANS and bis-ANS, may be employed in pharmacokinetic research. Although ANS and bis-ANS have become prominent fluorescent probes for protein characterization, the structural and spectroscopic characteristics of protein-ANS/bis-ANS complexes remain largely unknown. Both fluorescent dyes bind to either the folded or partially folded hydrophobic regions of proteins. Additionally, they serve to identify molten globule-like intermediates. These probes have been used to study the folding problems of protein structures and the mechanisms of protein-protein interactions. ANS and bis-ANS exhibited significant enhancement and blue shift in their emission spectra upon binding to TcArgRS, the primary enzyme responsible for attaching l-arginine to its corresponding tRNA. Through fluorescence spectroscopy and computational studies, we concluded that bis-ANS binds more tightly to TcArgRS and that ATP affects bis-ANS fluorescence signal. Thus, these probes are useful resources for studying the intricate intermolecular relationships between proteins in terms of their structure, function, and mechanism. Our study provides a framework for identifying the hydrophobic regions present in TcArgRS. The utilization of hydrophobic patches on proteins for drug targeting is noteworthy because they can assist in identifying regions on the surface of proteins that are likely to interact with ligands. These patches help identify hotspot residues that play a vital role in determining binding affinity. Drugs are mainly small and hydrophobic in nature, and they target protein surfaces which have complementary properties. In this study, we elucidated the potential of TcArgRS as a target for combating trypanosomal diseases and extending life expectancy.