Cloning and characterization of metallothionein gene (HcMT) from Halostachys caspica and its expression in E. coli.

Halostachys caspica is a short shrub distributed in the semi-arid and saline-alkali area, which evolved various mechanisms for modulating salt and metal level. In the present study, a Type 2 metallothionein (HcMT) gene was cloned from the salt induced suppression subtractive hybridization (SSH) cDNA library of H.caspica. Quantitative real time PCR (qRT-PCR) analysis indicated that HcMT gene was up-regulated under the stress of Cu(2+), Zn(2+) and Cd(2+), and the tolerance of E. coli strain harboring with the recombinant HcMT (pET-32a-HcMT) to Cu(2+), Zn(2+) and Cd(2+) was enhanced compared to strain with control vector (pET-32a). Moreover, the purified TrxA-HcMT fusion protein from E. coli cells grown in the presence of 0.3mM CuSO4, 0.3mM ZnSO4, or 0.1mM CdCl2 could bind more metal ions than TrxA alone. The predicted 3D structure showed that HcMT could form a single metal-thiolate cluster, which confers the ability to bind five divalent metal ions through fourteen cysteine residues. These data indicate that HcMT may be involved in processes of metal tolerance in H. caspica and could be employed as a potential candidate for heavy metal phytoremediation.