Department of Chemical Biology, Faculty of Biotechnology, University of Wrocław, F. Joliot-Curie 14a, 50-383 Wrocław, Poland.
Department of Chemistry and Biochemistry, Mendel University in Brno, Zemedelska 1, 613 00 Brno, Czech Republic; Central European Institute of Technology, Brno University of Technology, Purkynova 123, 612 00 Brno, Czech Republic.
Bioelectrochemistry. 2020 Aug;134:107501. doi: 10.1016/j.bioelechem.2020.107501. Epub 2020 Mar 14.
We focused on the application of mass spectrometry and electrochemical methods combined with a chemometric analysis for the characterization of partially metallothionein-3 species. The results showed decreased Cat1 and Cat2 signals for the Zn(II)-loaded MT3 species with respect to the metal-free protein, which might be explained by the arrangement of tetrahedral metal-thiolate coordination environments and the formation of metal clusters. Moreover, there was a decrease in the Cat1 and Cat2 signals, and a plateau was reached with 4-5 Zn(II) ions that corresponded to the formation of the C-terminal α-domain. Regarding the ZnMT3 complexes, we observed three different electrochemical behaviours for the ZnMT3, ZnMT3 and ZnMT3 species. The difference for ZnMT3 might be explained by the formation of independent ZnS cores in this stage that differ with respect to the formation of ZnCys clusters with an increased Zn(II) loading. The binding of the third Zn(II) ion to MT3 resulted in high sample heterogeneity due the co-existence of ZnMT3. Finally, the ZnMT3 protein showed a third type of behaviour. The fact that there were no free Cys residues might explain this phenomenon. Thus, this research identifies the major proteins responsible for zinc buffering in the cell.
我们专注于将质谱和电化学方法与化学计量分析相结合，用于部分金属硫蛋白-3 物种的表征。结果表明，与金属自由蛋白相比，Zn(II)负载的 MT3 物种的 Cat1 和 Cat2 信号降低，这可能是由于四面体金属-硫醇配位环境的排列和金属簇的形成。此外，Cat1 和 Cat2 信号降低，并在 4-5 个 Zn(II)离子处达到平台，对应于 C 端α-结构域的形成。关于 ZnMT3 配合物，我们观察到 ZnMT3、ZnMT3 和 ZnMT3 物种的三种不同电化学行为。对于 ZnMT3 的差异可能是由于在此阶段形成了独立的 ZnS 核，与形成具有增加 Zn(II)负载的 ZnCys 簇不同。第三个 Zn(II)离子与 MT3 的结合由于 ZnMT3 的共存导致样品高度不均匀。最后，ZnMT3 蛋白表现出第三种行为。没有游离 Cys 残基的事实可能解释了这一现象。因此，这项研究确定了细胞中锌缓冲的主要蛋白质。
