Identification and characterization of a recombinant metallothionein protein from a marine alga, Fucus vesiculosus

A cDNA library was constructed from macroalgae adapted to prolonged elevate d environmental copper levels. To investigate the possible existence of a m etallothionein (MT) gene, the library was screened with degenerate probes d esigned using plant MT cysteine-rich motifs. A gene was identified (1229 bp ) with a putative open reading frame (204 bp) encoding a 67-amino-acid prot ein exhibiting several characteristic features of MT proteins, including 16 cysteine residues (24%) and only one aromatic residue. Although the protei n sequence showed high identity with plant and invertebrate MTs, it contain ed a unique 'linker' region (14 amino acid residues) between the two putati ve metal-binding domains which contained no cysteine residues. This extende d linker is larger than the tripeptide found in archetypal vertebrate MTs, but does not conform either with the 40-amino-acid linkers commonly found i n plant MT sequences. An S-peptide Fucus MT fusion protein expressed in Esc herichia coli exhibited a relative molecular mass of similar to 14 kDa. The recombinant fusion bound seven Cd ions, of which 50% were dissociated at p H 4.1. Under anaerobic conditions, the Cd ions were displaced by Cu(I), whi ch associated with the protein at a ratio of 13 : 1. Laboratory exposure of F. vesiculosus to elevated copper resulted in induction of the MT gene. Th us this paper describes, for the first time, an MT gene identified from mac roalgae which is induced by copper exposure and whose encoded protein produ ct binds cadmium and copper.