Difference between revisions of "Part:BBa K391006"
(→Characterization Data) |
(→Characterization Data) |
||
| Line 3: | Line 3: | ||
== Characterization Data == | == Characterization Data == | ||
| − | The activity of DspB was tested by taking a crude cell lysate from <i>E.coli</i> cells expressing the protein. The protein in the lysate was then tested against a chromogenic substrate: 4-Nitrophenyl N-acetyl-β-D-glucosaminide. As DspB cleaves this substrate, the absorbance shifts from 300nm (substrate: 4-Nitrophenyl N-acetyl-β-D-glucosaminide) to 405nm (product: 4-nitrophenol). We measured the change in the level of absorbance at 405nm over time. | + | The activity of DspB was tested by taking a crude cell lysate from <i>E.coli</i> cells expressing the protein. The protein in the lysate was then tested against a chromogenic substrate: 4-Nitrophenyl N-acetyl-β-D-glucosaminide. As DspB cleaves this substrate, the absorbance shifts from 300nm (substrate: 4-Nitrophenyl N-acetyl-β-D-glucosaminide) to 405nm (product: 4-nitrophenol). We measured the change in the level of absorbance at 405nm over time and the result is shown below. |
<div style="text-align:center;">[[Image:Igem2010_assay2_linegraph.jpg|500px]]</div> | <div style="text-align:center;">[[Image:Igem2010_assay2_linegraph.jpg|500px]]</div> | ||
The graph above plots the absorbance at wavelength 405nm (y-axis) over time in days (x-axis). It can be observed from the graph that over time, the absorbance at 405nm increases with the treatment with the lysate containing DspB by a significant amount compared to the controls. This assay has been replicated and the results have shown to be reproducible. | The graph above plots the absorbance at wavelength 405nm (y-axis) over time in days (x-axis). It can be observed from the graph that over time, the absorbance at 405nm increases with the treatment with the lysate containing DspB by a significant amount compared to the controls. This assay has been replicated and the results have shown to be reproducible. | ||
Revision as of 05:55, 27 October 2010
DspB: carbohydrate digesting enzyme
Dispersin B (DspB) is an enzyme that degrades biofilms by catalyzing the hydrolysis of poly-ß-(1,6)-linked N-acetylglucosamine bonds. These bonds exist in the extracellular polymeric substance (EPS), which acts as a polysaccharide adhesin relevant to biofilm formation and integrity in Escherichia coli and Staphylococcus epidermidis. According to literature, DspB effectively cleaves these bonds and impedes biofilm formation.
Characterization Data
The activity of DspB was tested by taking a crude cell lysate from E.coli cells expressing the protein. The protein in the lysate was then tested against a chromogenic substrate: 4-Nitrophenyl N-acetyl-β-D-glucosaminide. As DspB cleaves this substrate, the absorbance shifts from 300nm (substrate: 4-Nitrophenyl N-acetyl-β-D-glucosaminide) to 405nm (product: 4-nitrophenol). We measured the change in the level of absorbance at 405nm over time and the result is shown below.
The graph above plots the absorbance at wavelength 405nm (y-axis) over time in days (x-axis). It can be observed from the graph that over time, the absorbance at 405nm increases with the treatment with the lysate containing DspB by a significant amount compared to the controls. This assay has been replicated and the results have shown to be reproducible.