Department of Biology, University of Naples Federico II, 80126 Naples, Italy.
Department of Precision Medicine, School of Medicine, University of Campania "Luigi Vanvitelli", via Luigi de Crecchio, 80138 Naples, Italy.
Molecules. 2020 Jul 19;25(14):3278. doi: 10.3390/molecules25143278.
Mercury (Hg) is a global environmental pollutant that affects human and ecosystem health. With the aim of exploring the Hg-induced protein modifications, intact human erythrocytes were exposed to HgCl (1-60 µM) and cytosolic and membrane proteins were analyzed by SDS-PAGE and AU-PAGE. A spectrofluorimetric assay for quantification of Reactive Oxygen Species (ROS) generation was also performed. Hg exposure induces alterations in the electrophoretic profile of cytosolic proteins with a significant decrease in the intensity of the hemoglobin monomer, associated with the appearance of a 64 kDa band, identified as a mercurized tetrameric form. This protein decreases with increasing HgCl concentrations and Hg-induced ROS formation. Moreover, it appears resistant to urea denaturation and it is only partially dissociated by exposure to dithiothreitol, likely due to additional protein-Hg interactions involved in aggregate formation. In addition, specific membrane proteins, including band 3 and cytoskeletal proteins 4.1 and 4.2, are affected by Hg-treatment. The findings reported provide new insights into the Hg-induced possible detrimental effects on erythrocyte physiology, mainly related to alterations in the oxygen binding capacity of hemoglobin as well as decreases in band 3-mediated anion exchange. Finally, modifications of cytoskeletal proteins 4.1 and 4.2 could contribute to the previously reported alteration in cell morphology.
汞（Hg）是一种全球性的环境污染物，会影响人类和生态系统的健康。本研究旨在探索汞诱导的蛋白质修饰，将完整的人红细胞暴露于 HgCl（1-60µM）中，并通过 SDS-PAGE 和 AU-PAGE 分析细胞质和膜蛋白。还进行了用于定量测定活性氧（ROS）生成的荧光分光光度法测定。Hg 暴露会引起细胞质蛋白电泳谱的改变，血红蛋白单体的强度显著降低，同时出现 64kDa 条带，鉴定为汞化的四聚体形式。这种蛋白质随着 HgCl 浓度的增加和 Hg 诱导的 ROS 形成而减少。此外，它似乎对尿素变性有抗性，并且仅部分通过暴露于二硫苏糖醇而解离，这可能是由于形成聚集体涉及到额外的蛋白质-Hg 相互作用。此外，特定的膜蛋白，包括带 3 和细胞骨架蛋白 4.1 和 4.2，也受到 Hg 处理的影响。本研究报告的结果为理解 Hg 对红细胞生理的可能有害影响提供了新的见解，主要与血红蛋白的氧结合能力的改变以及带 3 介导的阴离子交换的减少有关。最后，细胞骨架蛋白 4.1 和 4.2 的修饰可能导致先前报道的细胞形态改变。
