Covalent
modification of proteins by reactive pollutants/metabolites
might trigger various toxicities resulting from the disruption of
protein structures and/or functions, which is critical for understanding
the mechanism of pollutants-induced toxicity. However, this mechanism
has rarely been touched on due to the lack of a methodology. In this
research, the protein modification of bisphenol A (BPA) in rats was
characterized using a series of liquid chromatography–tandem
mass spectrometry (LC–MS) approaches. BPA-modified cysteine
(Cys1) was first released from proteins via enzymatic hydrolysis and
identified using LC–MS. Moreover, the positive correlation
between Cys1 and hepatotoxicity indicated the involvement of protein
modification in BPA toxicity. Then, in vitro incubation of BPA with
amino acids and protein confirmed that BPA could specifically modify
cysteine residues of proteins after bioactivation and provided four
additional modification patterns. Finally, 24 BPA-modified proteins
were identified from the liver of BPA-exposed rats using proteomic
analysis, and they were mainly enriched in oxidative stress-related
pathways. The modification on superoxide dismutases, catalase, and
glutathione S-transferases disrupted their enzymatic functions, leading
to oxidative damage. These results revealed that the covalent protein
modification is an unignorable factor for BPA hepatotoxicity. Moreover,
the workflow can be applied to identify protein adducts of other emerging
contaminants and possible risk.