Construct Design

Because the metallothionein proteins that bind to the heavy metals are located within the E. coli, we have constructed BioBricks containing heavy metal transport proteins that will translocate surrounding heavy metals into the cell. The high-affinity nickel transport protein nixA, originating from the bacteria Helicobacter pylori, imports nearby Ni(2+) ions into the cell ^[1]. Normally used by H. pylori to allow for urease activity ^[2], nixA also shows promise for purposes of bioaccumulation and remediation^[3].

The first nickel BioBrick BBa_K1460003 consists of the Anderson promoter, the nixA gene, and a terminator, allowing for the constitutive expression of the nixA gene and the accumulation of nickel within the cell. The second nickel BioBrick BBaK1460006 was constructed by inserting our metallothionein construct, the T7 promoter and GST-YMT metallothionein gene, downstream of our first construct. This allows for the simultaneous constitutive expression of nixA for nickel uptake and accumulation and the induced expression of metallothioneins. As metallothoineins inhibit cell growth, utilizing inducible metallothionein expression permits the bacteria to adequately grow before producing metallothioneins.