| In modern society, heavy metals are used extensively in a wide variety of industrial applications, and anthropogenic metal contamination continues to be a serious environmental and health hazard. Current cleanup methods have many shortcomings and the search continues for faster, more selective, and more cost-effective systems for pollution treatment. One area of growing interest is bioremediation, the utilization of biological processes, particularly microbial, for treatment of contaminated areas.; Previous studies have shown that co-expressing E. coli with a membrane transport system and intracellular metallothionein allows the cells to selectively hyperaccumulate heavy metal ions. In this work, two different strains, one designed with the L. plantarum manganese transporter MntA and the other with E. hirae copper transporter CopA, were examined. Bioaccumulation was studied under a variety of conditions, both to characterize and compare transporter activity and test the possible bioremediation applicability of these strains. While these bacteria do not display optimal metal accumulation parameters, they do have several advantages over existing techniques.; A major goal of bioremediation work is to improve upon the natural abilities of the microorganisms; understanding the mechanisms and residues responsible for ion binding and translocation is the first step in that path. To that end, mutational studies were carried out on CopA and MntA to examine two key regions involved in metal binding: the central transmembrane channel of MntA and the N-terminal heavy metal binding site of CopA. Analysis of MntA revealed that three amino acid residues, N554, N611, and D612, are probably responsible for high affinity cadmium binding, while several others may play roles in stabilizing the ion and maintaining proper side chain alignment. Results of the CopA studies revealed that the N-terminal metal binding motif is not essential for copper transport, but may have a role in increasing turnover number by acting as an initial metal binding site. |
