The simultaneously functions of Metallothioneins (MTs) are relied on their metalation mechanisms that can be divided into non-cooperative, weakly cooperative and strongly cooperative mechanisms. In this study, we recombinantly synthesized OsMTI-1b, N- and C-terminal Cys-rich regions as glutathione-S-transferase (GST)-fusion proteins in E. coli. In comparison with control strains (The E. coli cells containing pET41a without gene), transgenic E. coli cells showed more tolerance against Cd²⁺ and Zn²⁺. The recombinant GST-proteins were purified using affinity chromatography. According to in vitro assays, the recombinant proteins showed a higher binding ability to Cd²⁺ and Zn²⁺. However, the affinity of apo-proteins to Cu²⁺ ions were very low. The coordination of Cd²⁺ ions in OsMTI-1b demonstrates a strongly cooperative mechanism with a priority for the C-terminal Cys-rich region that indicates the detoxifying of heavy metals as main role of P1 subfamily of MTs. While the metalation with Zn²⁺ conformed to a weakly cooperative mechanism with a specificity to N-terminal Cys-rich region. It implies the specific function of OsMTI-1b is involved in zinc homeostasis. Nevertheless, a non-cooperative metalation mechanism was perceived for Cu²⁺ that suggests the fully metalation does not occur and OsMTI-1b cannot play a significant role in dealing with Cu²⁺ ions.

金属硫蛋白（MTs）的同时功能取决于它们的金属化机制，可以将其分为非合作机制，弱合作机制和强合作机制。在这项研究中，我们重组合成OsMTI-1B，N-和C-末端的Cys丰富区域作为谷胱甘肽-S-转移酶（GST） -融合蛋白在大肠杆菌（E.coli） 。与对照菌株（含有无基因的pET41a的大肠杆菌细胞）相比，转基因大肠杆菌细胞显示出对Cd ²⁺和Zn ^2+的更大耐受性。使用亲和色谱法纯化重组GST蛋白。根据体外测定，重组蛋白与Cd ^2+的结合能力更高。和Zn ²⁺。然而，脱辅基蛋白对Cu ²⁺离子的亲和力很低。OsMTI-1b中Cd ²⁺离子的配位证明了一种强合作机制，其中C端富含Cys的区域具有优先权，这表明重金属的解毒是MTs P1亚家族的主要作用。而Zn ²⁺的金属化符合弱协作机制，对N末端富含Cys的区域具有特异性。这表明OsMTI-1b的特定功能与锌稳态有关。然而，对于Cu ²⁺，存在一种非合作的金属化机制，这表明不会发生完全金属化，并且OsMTI-1b不能在处理Cu ^2+中起重要作用。 离子。