Part:BBa_K1526008:Experience

Complementation assay for BW25113 (WT and Mutants for sulfate transporter).

In E. coli the main sulfate transporter is an ABC transporter and not a secondary passive transporter as CysP from Bacillus subtilis, therefore, our team has complemented mutants of BW25113 for different subunits of this permease (CysA - ATPase subunit - and CysW - permease subunit - mutants). We have shown that our gene has successfully complemented our mutants by growing our strains in M9 minimal medium supplemented with 0.4% glucose, 1mM of Magnesium Sulfate and 1mM of IPTG in order to induce our construct.

As seen in the graphic below, after 16 hours of growth in M9 our complemented mutants have reached almost the same growth as our Wild-Type strain. The wild-type strain as well has increased its growth after transformation with our CysP construct.

Applications of BBa_K1526008

As part of our final construct, CysP is responsible for increasing the influx of sulfates inside of the cell, increasing the cysteine production intracellularly. Therefore, the outcomes of this Biobrick are both removing sulfates from environment (that can be harmful in high concentrations) and at the same time targeting them to central metabolism in the cell for cysteine production.

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