HPLC characterization of Ag+ and Cu+ metal exchange reactions with Zn- and Cd-metallothioneins.
HPLC characterization of Ag+ and Cu+ metal exchange reactions with Zn- and Cd-metallothioneins.
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DOI：
10.1021/bi961402f
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发表时间：
1996-11
期刊：
影响因子：
2.9
通讯作者：
H. Li;J. Otvos
中科院分区：
文献类型：
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作者：
H. Li;J. Otvos
采用阴离子交换高效液相色谱法研究了ZnMT和CdMT与Ag+和Cu+的金属交换反应。这些反应的动力学揭示了存在一个缓慢的蛋白质间金属再分配过程，随后由Ag+或Cu+的金属从Zn 7 MT或Cd 7 MT的初始快速位移。在Chelex树脂存在下，Ag+与Zn 7 MT反应的动力学有利产物包括Zn 4Ag 6 MT和Ag 12 MT。随后Zn 7 MT和Ag 12 MT之间的缓慢反应导致了化学上有利的产物Zn 4Ag 6 MT。用Ag+滴定Cd 7 MT和用Cu+滴定Zn 7 MT，观察到类似的行为。发现使用Chelex树脂去除未结合的金属离子影响Zn 7 MT和Cd 7 MT与Ag+的反应。在不存在Chelex的情况下将Ag+添加到Zn 7 MT后，除了Zn 4 Ag 6 MT和Ag 12 MT之外，Ag 18 MT物质形成为动力学有利的产物。随后，发现Ag 18 MT通过缓慢的蛋白质间金属交换反应将Ag+提供给Zn 7 MT和Zn 4 Ag 6 MT。Chelex树脂通过限制Ag 18 MT的形成和/或通过结合带正电荷的Ag 18 MT物质并将其从溶液中除去来影响滴定反应。在不存在Chelex的情况下Ag 18 MT的形成解释了为什么需要超过12当量的Ag+来从Zn 7 MT和Cd 7 MT中置换所有的Zn 2+或Cd 2+。在这项研究中开发的HPLC方法允许均匀的制剂，由金属混合MT物种含有Ag+在β-域和Cd 2+或Zn 2+在α-域。
Anion exchange HPLC methods were developed to study the metal exchange reactions of ZnMT and CdMT with Ag+ and Cu+. The kinetics of these reactions revealed the existence of a slow interprotein metal redistribution process that follows initial rapid displacement of metal from Zn7MT or Cd7MT by Ag+ or Cu+. Kinetically-favored products of the reaction of Ag+ with Zn7MT in the presence of Chelex resin include Zn4Ag6MT and Ag12MT. Subsequent slow reaction between Zn7MT and Ag12MT leads to the thermodynamically-favored product, Zn4Ag6MT. Analogous behavior was observed for Cd7MT titrated with Ag+ and Zn7MT with Cu+. The use of Chelex resin to remove unbound metal ions was found to influence the reactions of Zn7MT and Cd7MT with Ag+. Upon addition of Ag+ to Zn7MT in the absence of Chelex, a Ag18MT species forms as the kinetically-favored product in addition to Zn4Ag6MT and Ag12MT. Subsequently, Ag18MT was found to donate Ag+ to Zn7MT and Zn4Ag6MT via slow interprotein metal exchange reactions. Chelex resin influences the titration reaction either by limiting formation of Ag18MT and/or by binding the positively charged Ag18MT species and removing it from solution. The formation of Ag18MT in the absence of Chelex explains why more than 12 equiv of Ag+ is required to displace all of the Zn2+ or Cd2+ from Zn7MT and Cd7MT. The HPLC methods developed in this study allow homogeneous preparations to be made of metal-hybrid MT species containing Ag+ in the beta-domain and Cd2+ or Zn2+ in the alpha-domain.