Identifying Novel Membrane Proteins for Bioproduction

Model microbes such as bacteria and yeast are often used for bioproduction of fuels and chemicals. Target molecules can be toxic to these microbes, but this limitation can be overcome by using membrane proteins that confer resistance to toxic compounds. Small Multidrug Resistance transporters (SMRs) are proteins that function to expel drug-like substances from cells and contribute to bacterial antibiotic resistance. We identified novel putative SMRs in non-model anaerobic fungi, including Neocallimastix californiae. This project aims to determine if N. californiae SMRs can function in the model organisms Escherichia coli and yeast, Saccharomyces cerevisiae. The SMR gene was cloned in plasmid vectors, tagged with a Green Fluorescent Protein (GFP) reporter, and transformed into E. coli and S. cerevisiae. Expression of the SMR gene was detected in both E. coli and S. cerevisiae using SDS-PAGE in-gel GFP fluorescence. In future work, tolerance assays will be used to investigate if the SMR increases the tolerance of model organisms to various drug-like compounds. A fungal SMR functioning in E. coli or yeast could provide insight into how to genetically engineer model microbes for more efficient bioproduction.