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Results obtained from rat studies indicate that, even at low concentrations, mercurial species cause harmful effects on the kidneys, by inducing the nephrotic oxidative stress response. In the present work, Hg-associated proteins were identified as possible mercury-exposure biomarkers in rat kidneys exposed to low mercury chloride concentrations for 30 days (Hg-30) and 60 days (Hg-60), using metalloproteomic strategies. The renal proteomic profile was fractioned by two-dimensional electrophoresis and the mercury determinations in kidney samples, protein pellets and protein spots were performed using graphite furnace atomic absorption spectrometry. The characterization of Hg-associated protein spots and the analysis of differentially expressed proteins were performed by liquid chromatography, coupled with tandem mass spectrometry. Eleven Hg-associated protein spots with a concentration range of 79 ± 1 to 750 ± 9 mg kg -1 in the Hg-60 group were identified. The characterization and expression analyses allowed the identification of 53 proteins that were expressed only in the Hg-60 group, 13 "upregulated" proteins ( p > 0.95) and 47 "downregulated" proteins ( p < 0.05). Actin isoforms and hemoglobin subunits were identified in protein spots of the Hg-60 group, with mercury concentrations in the range of 138 to 750 mg kg -1 , which qualifies these proteins as potential mercury-exposure biomarkers., Competing Interests: The authors declare no conflict of interest.

The aims of this study were to identify mercury-associated protein spots in the liver tissue of rats exposed to low concentrations of mercury and to elucidate the physiological and functional aspects of the proteins identified in the protein spots. Therefore, proteomic analysis of the liver tissue of Wistar rats exposed to mercury chloride (4.60 μg kg -1 in Hg 2+ ) was performed for thirty days (Hg-30 group) and sixty days (Hg-60 group). The proteomic profile of the liver tissue of the rats was obtained by two-dimensional electrophoresis (2D-PAGE), and the determinations of total mercury in the liver tissue, pellets and protein spots were performed by graphite furnace atomic absorption spectrometry (GFAAS). ImageMaster 2D Platinum 7.0 software was used to identify the differentially expressed mercury-associated protein spots, which were then characterized by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). The determinations by GFAAS indicated a total mercury bioaccumulation of 2812% in the Hg-30 group and 3298% in the Hg-60 group and 10 mercury-associated protein spots with a concentration range of 51 ± 1.0 to 412 ± 6.00 mg kg -1 in the 2D PAGE gels from the liver tissue of the Hg-60 group. The LC-MS/MS analyses allowed the identification of 11 metal binding proteins in mercury-associated protein spots that presented fold change with upregulation >1.5, downregulation < -1.7 or that were expressed only in the Hg-60 group. Using the FASTA sequences of the proteins identified in the mercury-associated protein spots, bioinformatics analyses were performed to elucidate the physiological and functional aspects of the metal binding proteins, allowing us to infer that enzymes such as GSTM2 presented greater mercury concentrations and downregulation < -3; Acaa2 and Bhmt, which showed expression only in the Hg-60 group, among others, may act as potential mercury exposure biomarkers., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)