Difference between revisions of "Part:BBa K314012"
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To test BBa _K314012, it was inserted into a pET29b+ vector using Kunkel's Mutagenesis. CapD_CP was then produced and purified as described in the [http://2010.igem.org/Team:Washington/Protocols/50mLPurificationCapD UW 2010 iGEM Team's Small Scale Protein Purification Protocol]. The purified protein was then tested for activity. For a detailed description of the assay, please see the [http://2010.igem.org/Team:Washington/Protocols/EnzymeAssayCapD 2010 UW iGEM wiki]. The resulting data is shown below. | To test BBa _K314012, it was inserted into a pET29b+ vector using Kunkel's Mutagenesis. CapD_CP was then produced and purified as described in the [http://2010.igem.org/Team:Washington/Protocols/50mLPurificationCapD UW 2010 iGEM Team's Small Scale Protein Purification Protocol]. The purified protein was then tested for activity. For a detailed description of the assay, please see the [http://2010.igem.org/Team:Washington/Protocols/EnzymeAssayCapD 2010 UW iGEM wiki]. The resulting data is shown below. | ||
| − | <gallery heights= | + | <gallery heights=350px widths=350> |
image:CapDCP_MM.png | image:CapDCP_MM.png | ||
image:CapDandCapDCPGel.jpg | image:CapDandCapDCPGel.jpg | ||
Revision as of 17:51, 22 October 2010
CapD_CP
A circularly permuted [http://www.parts.igem.org/Part:BBa_K314011 CapD] with a Foldit-designed linker.
To test BBa _K314012, it was inserted into a pET29b+ vector using Kunkel's Mutagenesis. CapD_CP was then produced and purified as described in the [http://2010.igem.org/Team:Washington/Protocols/50mLPurificationCapD UW 2010 iGEM Team's Small Scale Protein Purification Protocol]. The purified protein was then tested for activity. For a detailed description of the assay, please see the [http://2010.igem.org/Team:Washington/Protocols/EnzymeAssayCapD 2010 UW iGEM wiki]. The resulting data is shown below.
The substrate vs. reaction rate curve above plots the PDG cleaved per enzyme per hour (y-axis) as a function of substrate concentration (x-axis). As observed in the curve above, high substrate concentrations suffered from substrate inhibition under the conditions our enzyme was assayed. Kinetic constants, kcat and Km, taken from our plotted curves are shown in the table above.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 1489
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]