Harnessing the BrkA Autotransporter to Surface Display the Zinc-Binding Domain Tetrahymena thermophila Metallothionein MTT5 on Escherichia coli for Zinc Biosorption Evaluation

Heavy metal contamination, particularly zinc, poses significant environmental and health concerns. Surface expression of metal-binding proteins in easily culturable bacteria such as Escherichia coli presents a promising bioremediation strategy to mitigate heavy metal pollution. We investigated the surface expression of Tetrahymena thermophila metallothionein (MTT5), a 10.5 kDa metal-binding protein, using the BrkA autotransporter system in E. coli. We designed a recombinant BrkA-MTT5 plasmid (pADKT5) and confirmed MTT5 expression via western blot analysis. Further investigation utilizing a trypsin accessibility assay suggested the potential for BrkA-mediated export of MTT5. Additionally, the zinc-binding capacity of BrkA-MTT5 was evaluated by an Arabidopsis thaliana root growth assay which demonstrated statistically significant differences between bacteria-treated and non-treated conditions. Overall, these findings highlight the potential for BrkA-mediated export of MTT5 and lay the foundation for further research aimed at harnessing the bacterial surface display system for heavy metal bioremediation.