A mass spectrometry-based approach gives new insight into organotin-protein interactions.

Abstract

In this study, the combination of speciation analysis and native mass spectrometry is presented as a powerful tool to gain new insight into the diverse interactions of environmentally relevant organotin compounds (OTCs) with proteins. Analytical standards of model proteins, such as β-lactoglobulin A (LGA), were thereby incubated with different phenyl- and butyltins. For adduct identification and characterization, the incubated samples were analyzed by inductively coupled plasma-mass spectrometry (ICP-MS) and electrospray ionization-mass spectrometry (ESI-MS) in combination with size exclusion chromatography (SEC). It allowed for a mild separation, which was most crucial to preserve the acid-labile organotin-protein adducts during their analyses. The binding of triorganotin compounds, such as triphenyltin, was shown to be sulfhydryl-directed by using cysteine-specific protein labeling. However, the sole availability of reduced cysteine residues in proteins did not automatically enable adduct formation. This observation complements previous studies and indicates the necessity of a highly specific binding pocket, which was identified for the model protein LGA via enzymatic digestion experiments. In contrast to triorganotins, their natural di- and mono-substituted degradation products, such as dibutyltin, revealed to be less specific regarding their binding to several proteins. Further, it also did not depend on reduced cysteine residues within the protein. In this context, they can probably act as linker molecules, interconnecting proteins, and leading to dimers and probably to higher oligomers. Furthermore, dibutyltin was observed to induce hydrolysis of the protein's peptide backbone at a specific site. Concerning unknown long-term toxic effects, our studies emphasize the importance of future studies on di- and mono-substituted OTCs.