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− | <p class = "caption"><b>Figure | + | <p class = "caption"><b>Figure 3:</b> Figure 3: chrR6 reductase enzyme is more efficient than nemA reductase in reducing Cr(VI) </p> |
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<h2>cysPUWA Effect on Cell Membrane Permeability to Cr(VI)</h2> | <h2>cysPUWA Effect on Cell Membrane Permeability to Cr(VI)</h2> | ||
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+ | <p> n this experiment, we added varying concentrations of Cr(VI) to 0.05 OD<sub>600</sub> cultures of: <i>E. coli</i> MG1655, <i>E. coli</i> MG1655 transformed with a plasmid constitutively expressing reductase chrR6, and <i>E. coli</i> MG1655 cotransformed with a plasmid constitutively expressing reductase chrR6 and the sulfate transporter system cysPUWA-sbp. We grew the cultures for 12 hours shaking at 37C and took their final OD<sub>600</sub> measurements (presented here). The data indicates that the sulfate transporter system introduces toxicity to the cell (because it has the lowest OD<sub>600</sub> values for every initial concentration of Cr(VI) except [Cr(VI)]<sub>i</sub>=0). Because the OD<sub>600</sub> values of the reductase and co-transformed reductase/transporter cultures are comparable at time 12 hours for no chromate, this indicates that the additional toxicity from the sulfate transporters is due to increase permeability of the cell to Cr(VI) rather than increased metabolic load of expressing a sythetically engineered plasmid. </p> | ||
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+ | <img src="https://static.igem.org/mediawiki/parts/d/d4/Corrected_c6%2C_cys.png" " style="width:60%"> | ||
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+ | <p class = "caption"><b>Figure 3:</b> Figure XXX: chrR6 reductase enzyme is more efficient than nemA reductase in reducing Cr(VI) </p> | ||
+ | </img> | ||
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Revision as of 03:42, 2 November 2017
RESULTS
Overview
chrR6 Chromium Reduction Activity
As detailed on our "Experiments" page, we measured chrR6 chromium reduction activity using DPC assay. The strong r2 value of 0.9944 for the linear trend line of our standard DPC curve indicates that this is an accurate Cr(VI) detection method within this range of concentrations. Therefore, when we investigate the reduction efficiencies of our reductases, we will use initial Cr(VI) concentrations within this range.
Figure 2.A compares the Cr(VI) reduction rate standardized for OD600 of E. coli MG1655 versus E. coli MG1655 cotransformed with the sulfate transporter system (BBa_K2194004) and constitutively expressed chrR6 reductase enzyme (BBa_K2194000). The data indicates that below an initial [Cr(VI)] of 80 uM the cotransformed bacteria are more efficient at reducing Cr(VI) than wild type bacteria. Above 80 uM initial [Cr(VI)], the trend reverses and wild type bacteria are more efficient. This supports the hypothesis that increasing cell permeability to chromate (in this case via sulfate transporters) increases the reduction efficiency of chrR6, but that after a threshold concentration the increased permeability becomes toxic for the cell.