Metal binding ability of metallothionein-3 expressed in Escherichia coli

Metallothionein-3/growth inhibitory factor is a brain-specific member of the metallothionein gene family, and impairs the survival and neurite formation of cultured neurons. Metallothionein-3 can bind heavy metals such as Zn, Cu, or Cd almost in the same way as other metallothionein family. However, its biological function as growth inhibitory factor apparently distinguishes metallothionein-3 from other metallothioneins. To better understanding of the relationship between the growth inhibitory activity of metallothionein-3 and metals bound to metallothionein-3, the metal-binding ability of metallothionein-3 was analyzed in comparison with those of metallothionein-1 and -2. The metal-binding ability of metallothionein-3 was evaluated by pH titration and 5-5' dithiobis (2-nitrobenzoic) acid (DTNB) analysis as compared with those of the other metallothioneins. The affinity of metal ions for metallothionein-3 was indicated as follows, Cu>Cd>Zn, same as metallothionein-1 and -2. However, the affinity of metallothionein-3 to Cu was much higher than that of metallothionein-1 and -2. The strong affinity to Cu of metallothionein-3 might be related to its growth inhibitory activity.