On the sensitivity of metallothioneins to oxidation during isolation.
On the sensitivity of metallothioneins to oxidation during isolation.
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关于金属硫蛋白在分离过程中对氧化的敏感性。
DOI：
10.1042/bj1910475
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发表时间：
1980
期刊：
影响因子：
--
通讯作者：
D. Petering
中科院分区：
文献类型：
--
作者：
D. Minkel;K. Poulsen;S. Wielgus;C. Shaw;D. Petering
结果表明，暴露于氧化条件下，金属在大鼠肝和马肾上清液Sephadex G-75组分中的分布发生改变。特别是，在有氧条件下，与金属硫蛋白结合的金属被置换成高分子量的组份，在较小程度上被置换成低分子量的形式。在这个过程中，金属硫蛋白锌比镉更不稳定。在高相对分子质量组分中也发现了相当大比例的硫蛋白，并可以通过硫醇处理与金属一起回收。这一结果表明，在有氧条件下获得的高相对分子质量组分中镉含量较高的分布是金属硫蛋白氧化造成的假象，并表明镉等金属的“溢出”可能在很大程度上是由于氧化过程而不是饱和效应。有证据表明，当硫蛋白结合在高分子量区域时，二硫键的形成就会发生，而化学还原是恢复其正常洗脱行为所必需的。加入到匀浆中的巯基乙醇保持了细胞环境中通常存在的还原条件，并防止了金属硫蛋白的氧化，在分离过程中金属重新分布。在这种情况下，从长期接触饮用水中低浓度镉的动物中分离出来的大鼠肝脏金属硫蛋白含有相当数量的铜和锌，并含有大量存在于马肾上清液中的锌。4万克球团经超声处理后也可提取金属硫蛋白。因此，对大鼠肝脏中镉沉积位置的仔细分析研究表明，超过95%的镉与金属硫蛋白结合。
It is demonstrated that the distribution of metals among the Sephadex G-75 fractions of rat liver and horse kidney supernatant is altered by exposure to oxidizing conditions. In particular, the metals bound to metallothionein are displaced into high-molecular-weight fractions and, to a lesser extent, into the low-molecular-weight forms, under aerobic conditions. In this process, metallothionein zinc is much more labile than cadmium. An appreciable proportion of the thionein is also found in the high-molecular-weight fractions and can be recovered along with the metals by treatment with mercaptoethanol. This result shows that the distributions obtained aerobically with large cadmium content in the high-molecular-weight fractions are an artefact due to metallothionein oxidation and suggests that 'spillage' of metals such as cadmium may be due in large part to oxidative processes rather than saturation effects. Evidence is presented that disulphide-bond formation occurs as thionein becomes bound in the high-molecular-weight region and that chemical reduction is necessary to restore its normal elution behaviour. Mercaptoethanol added to the homogenates maintains the reducing conditions normally found in the cellular milieu and prevents the oxidation of the metallothionein redistribution of the metals during isolation. Under these conditions the rat liver metallothionein isolated from animals exposed to chronic low concentrations of cadmium in drinking water contains appreciable quantities of copper as well as zinc and contains much of the zinc that is present in horse kidney supernatants. Metallothionein can also be extracted from a 40 000g pellet after sonication of the pellet. Thus careful analytical studies of the sites of cadmium deposition in rat liver indicate that greater than 95% is bound to metallothionein.