Production and cellular localization of functional oligomeric peptides in E. coli: expression of the N. crassa polymetallothionein

Tandem repeat oligomers of the Neurospora crassa metallothionein peptide were expressed in Escherichia coli and targeted to the periplasm as a fusion to the maltose binding protein. Production of these peptide oligomers in E. coli resulted in enhanced cadmium removal from solutions of cells containing short number of repeats. However, cells containing longer peptide oligomers demonstrated diminished cadmium removal capacity. Cellular distribution of expressed fusion protein revealed that while the majority of the short-tandem oligomers reside within the periplasmic space, cells expressing the longer oligomers harbor these proteins as insoluble inclusion bodies that are not localized properly. Analysis of the purified proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed a ladder of protein bands indicating the possibility of proteolysis or premature translation termination of the repeating peptide oligomers. These results indicate that although metal removal by cells can be enhanced through production of tandem repeats, due to the nature of the repetitive sequences, or other factors, longer oligomers do not properly localize within the periplasmic space, and therefore a diminished rate of metal accumulation is observed by these cells.