Difference between revisions of "Part:BBa K3843002"
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1UTM-HRP is a translational fusion of Atlantic salmon trypsin (1UTM) and horseradish peroxidase (HRP). 1UTM can bind to PEA (a natural inhibitor of salmon trypsin), and HRP is a chemiluminescent enzyme. A 6-His tag followed by a TEV protease linker was added at the N-terminus to allow for purification by Ni-NTA affinity chromatography. At the C-terminus, an Avi-tag was appended, allowing for biotinylation. A flexible, moderately long (13 AA), and polar linker (consisting of Ser and Thr) separated 1UTM and HRP, allowing for proper functioning and stability of the fusion protein in aqueous solution. | 1UTM-HRP is a translational fusion of Atlantic salmon trypsin (1UTM) and horseradish peroxidase (HRP). 1UTM can bind to PEA (a natural inhibitor of salmon trypsin), and HRP is a chemiluminescent enzyme. A 6-His tag followed by a TEV protease linker was added at the N-terminus to allow for purification by Ni-NTA affinity chromatography. At the C-terminus, an Avi-tag was appended, allowing for biotinylation. A flexible, moderately long (13 AA), and polar linker (consisting of Ser and Thr) separated 1UTM and HRP, allowing for proper functioning and stability of the fusion protein in aqueous solution. | ||
The resulting fusion protein can be used for the detection of PEA in a diagnostic microfluidic assay that produces a quantifiable chemiluminescent signal, which was the basis of Waterloo iGEM 2021's project. | The resulting fusion protein can be used for the detection of PEA in a diagnostic microfluidic assay that produces a quantifiable chemiluminescent signal, which was the basis of Waterloo iGEM 2021's project. | ||
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The following image depicts the 1UTM-HRP fusion (with 1UTM on the left and HRP on the right). The 3D model was obtained by multi-domain structural modelling on UCSF Chimera (Pettersen et al., 2004). | The following image depicts the 1UTM-HRP fusion (with 1UTM on the left and HRP on the right). The 3D model was obtained by multi-domain structural modelling on UCSF Chimera (Pettersen et al., 2004). | ||
Revision as of 18:02, 18 October 2021
1UTM-HRP (fusion)
1UTM-HRP is a translational fusion of Atlantic salmon trypsin (1UTM) and horseradish peroxidase (HRP). 1UTM can bind to PEA (a natural inhibitor of salmon trypsin), and HRP is a chemiluminescent enzyme. A 6-His tag followed by a TEV protease linker was added at the N-terminus to allow for purification by Ni-NTA affinity chromatography. At the C-terminus, an Avi-tag was appended, allowing for biotinylation. A flexible, moderately long (13 AA), and polar linker (consisting of Ser and Thr) separated 1UTM and HRP, allowing for proper functioning and stability of the fusion protein in aqueous solution. The resulting fusion protein can be used for the detection of PEA in a diagnostic microfluidic assay that produces a quantifiable chemiluminescent signal, which was the basis of Waterloo iGEM 2021's project.
The following image depicts the 1UTM-HRP fusion (with 1UTM on the left and HRP on the right). The 3D model was obtained by multi-domain structural modelling on UCSF Chimera (Pettersen et al., 2004).
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1113
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 1077
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 1249