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The toxicity of lead, one of the most ubiquitous toxic metals, is well known. Some of its pathological effects are related to its preference for the sulfhydryl groups of proteins. Metallothioneins (MT) are a particular family of metalloproteins characterized by their high Cys content that, among other functions, are linked to the detoxification of heavy metals. In mammals, 4 MT isoforms have been found. The MT3 isoform, also called “neuronal growth inhibitory factor”, is mainly synthesized in the brain and contains several structural differences that may contribute to important functional differences between it and other MT isoforms. The abilities of recombinant MT3 and its individual αMT3 and βMT3 fragments to bind Pb( ii ) have been investigated here, under different pH conditions, by means of spectroscopy, mass spectrometry and isothermal titration calorimetry. The results obtained show that the binding of Pb( ii ) to the intact MT3 protein is relatively unaffected by pH, while the individual domains interact with Pb( ii ) in a pH-sensitive manner. The mass spectrometry data reveal the evolution with time of the initially formed Pb-MT complexes. In the case of the full length protein, Pb( ii ) remains bound for a long period of time. With the isolated fragments, the lead is eventually released. The Pb-species formed depend on the amount of Pb( ii ) present in solution. The thermodynamic data recorded, as measured by ITC, for the replacement of Zn( ii ) by Pb( ii ) in reactions with Zn-MT3, Zn-αMT3 and Zn-βMT3 are all similar, and in all cases, the displacement of Zn( ii ) by Pb( ii ) is thermodynamically favorable. Zn-Replete and Pb-replete MT3 have distinctive circular dichroism spectra, suggestive of structural differences with different metallation status. 

Abstract Zinc is important in neurogenesis, but excessive levels can cause apoptosis and other pathologies leading to cognitive impairments. Mast cells are present in many brain regions including the hippocampus, an area rich in vesicular zinc. Mast cells contain zinc‐rich granules and a well‐developed mechanism for uptake of zinc ions; both features point to the potential for a role in zinc homeostasis. Prior work using the Timm stain supported this hypothesis, as increased labile zinc was detected in the hippocampus of mast cell‐deficient mice compared to wild‐type mice while no differences in total zinc were found between the two genotypes in the whole brain or other tissues. The current report further examines differences in zinc homeostasis between wild‐type and mast cell‐deficient mice by exploring the zinc transporter ZnT3, which transports labile zinc into synaptic vesicles. The first study used immunocytochemistry to localize ZnT3 within the mossy fibre layer of the hippocampus to determine whether there was differential expression of ZnT3 in wild‐type versus mast cell‐deficient mice. The second study used inductively coupled plasma mass spectrometry (ICP‐MS) to determine total zinc content in the whole dentate gyrus of the two genotypes. The immunocytochemical results indicate that there are higher levels of ZnT3 localized to the mossy fibre layer of the dentate gyrus of mast cell‐deficient mice than in wild‐type mice. TheICP‐MSdata reveal no differences in total zinc in dentate gyrus as a whole. The results are consistent with the hypothesis that mast cells participate in zinc homeostasis at the level of synaptic vesicles.