Analysis of mammalian metallothionein isoforms by high-resolution SDS-gel electrophoresis

Metallothioneins (MTs) are cysteine-rich heavy metal-binding proteins, whose possible functions are thought to be the protection against toxic metals as well as the regulation of essential metals. It is known that there are several MT isoforms, but the biological roles of the individual isoforms have not been elucidated. To facilitate the functional analysis of these isoforms, we improved an analytical method of MTs developed previously, which is based on a denaturing gel electrophoresis of chemically modified MTs. The established technique makes it possible not only to separate MT isoforms with a high resolution, but to estimate the levels of the individual isoforms by analyzing directly crude cell extracts. By this method, six MT isoforms were identified in the extracts of Cd-exposed human cells. It was also revealed that there is an apparent heterogeneity of the rat liver MT; five isoforms were identified in the liver extracts of Cd-injected rats. The present method will be useful in the functional analysis of the MT isoforms, as well as in a variety of aspects of the MT studies.