Difference between revisions of "Part:BBa K1936001"

Revision as of 22:44, 15 October 2016

Tom5-eGFP-LOV2

ALERT !!!Under construction !!

Usage

While LOV2 domain was previously described for optogenetical control of gene expression (BBa_K1742000) and as reporter gene (BBa_K660003), we took a new approach. The described fusion protein Tom5-eGFP-LOV2 is designed to recruit proteins of choice to the outer mitochondrial membrane (OMM) in engineered eukaryotic cells.

Tom5 is a part of the eukaryotic translocase of outer membrane protein complex and has a single membrane anchor unit that integrates into OMM. [1] We fused it to the N-terminal end of LOV2 domain from A.sativa. Upon exposure to blue light (λ=473nm) the LOV2 peptide undergoes a conformational change and exposes its C-terminal J-alpha-helix which is then able to bind a specifically engineered PDZ domain (BBa_K1470005). [2]

The fused enhanced green fluorenscence protein (eGFP) is used to track protein localization by microscopy.

Our blue light dependent system can be used to recruit any protein of interest to the outer mitochondrial membrane by simply fusing it to an ePDZ domain.

: Figure 2: The LOV2-based optogenetic switch is activated by blue light.

Biology

Characterization

References

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 33
Illegal BamHI site found at 712
Illegal XhoI site found at 515
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 1226

@@ Line 8: / Line 8: @@
 ===Usage===
-While LOV2 domain was previously described for optogenetical control of gene expression (BBa_K1742000) and promotor characterization (BBa_K660004), we took a new approach. The described fusion protein Tom5-eGFP-LOV2 is designed to recruit proteins of choice to the outer mitochondrial membrane (OMM) in engineered eukaryotic cells.
+While LOV2 domain was previously described for optogenetical control of gene expression (<html><a href="https://parts.igem.org/Part:BBa_K1742000">BBa_K1742000</a></html>) and as reporter gene (<html><a href=" https://parts.igem.org/Part:BBa_K660003">BBa_K660003</a></html>), we took a new approach. The described fusion protein Tom5-eGFP-LOV2 is designed to recruit proteins of choice to the outer mitochondrial membrane (OMM) in engineered eukaryotic cells.
-Tom5 is a part of the eukaryotic translocase of outer membrane protein complex and has a single membrane anchor unit that integrates into OMM. [1] We fused it to the N-terminal end of LOV2 domain from <html><i>A.sativa</i></html>. Upon exposure to blue light (473nm) the LOV2 peptide undergoes a conformational change and exposes its C-terminal J-alpha-helix which is then able to bind a specifically engineered PDZ domain (BBa_K1470005). [2]
+Tom5 is a part of the eukaryotic translocase of outer membrane protein complex and has a single membrane anchor unit that integrates into OMM. [1] We fused it to the N-terminal end of LOV2 domain from <html><i>A.sativa</i></html>. Upon exposure to blue light (<html>&lambda;</html>=473nm) the LOV2 peptide undergoes a conformational change and exposes its C-terminal J-alpha-helix which is then able to bind a specifically engineered PDZ domain (<html><a href="https://parts.igem.org/Part:BBa_K1470005">BBa_K1470005</a></html>). [2]
-Our blue light dependent system can be used to recruit any protein of interest to the outer mitochondrial membrane by simply fusing it to an ePDZ domain.
+The fused enhanced green fluorenscence protein (eGFP) is used to track protein localization by microscopy.
+Our blue light dependent system can be used to recruit any protein of interest to the outer mitochondrial membrane by simply fusing it to an ePDZ domain.