Difference between revisions of "Part:BBa K909007"

Revision as of 17:00, 26 September 2012

TetR DNA binding domain containing BamHI site for protein fusions

Truncated version of tetracycline repressor TetR, consisting of 1 – 127 amino acids, containing DNA binding domain. The rest of the protein, alpha 8-10 helices responsible for tetR dimerization. Without dimerization domain, tetR-DBD fails to bind TetR responsive promoter efficiently and unable to repress transcription of reporter protein (see figure). However, TetR-DBD fusion with UVR8 protein (BBa_K909008) restores TetR-DBD promoter binding.

GFP repression (reporter) by full length TetR, TetR-DBD and dimerizing TetR-DBD-UVR8 fusion

Usage and Biology

We introduced BamHI site at C terminus of tetR-DBD for an easy in frame fusions with other proteins. This can easily be used as a two hybrid system to detect homo/heterodimers in E. coli, and, in principle, one can use these fusions to turn protein-protein interaction into the repression of transcription. Furthermore, inducible dimerization can lead to a novel switch like behaviors of the system (e.g. [http://2012.igem.org/Team:ETH_Zurich ETH Zurich 2012 team] created a novel UV-B inducible-ON switch).

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 382
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

@@ Line 3: / Line 3: @@
-Truncated version of tetracycline repressor TetR, consisting of 1 – 127 amino acids, containing DNA binding domain. The rest of the protein, alpha 8-10 helixes responsible for tetR dimerization, were removed in order to keep tetR-DBD in monomer form and prevent from efficient DNA binding and transcription repression (see figure).
+Truncated version of tetracycline repressor TetR, consisting of 1 – 127 amino acids, containing DNA binding domain. The rest of the protein, alpha 8-10 helices responsible for tetR dimerization. Without dimerization domain, tetR-DBD fails to bind TetR responsive promoter efficiently and unable to repress transcription of reporter protein (see figure).  However, TetR-DBD fusion with UVR8 protein (<partinfo>BBa_K909008</partinfo>) restores TetR-DBD promoter binding.
-Fusions of TetR-DBD with a dimerizing protein(e.g. UVR8 (link))restores TetR-DBD DNA binding.
@@ Line 14: / Line 14: @@
 ===Usage and Biology===
-We introduced BamHI site at C terminus for an easy in tetR-DBD frame fusions with other proteins. Thus, it can be used in two hybrid systems for both, homo and hetero dimerizations, also one can use these fusions to turn protein-protein interaction into the repression of transcription.
+We introduced BamHI site at C terminus of tetR-DBD for an easy in frame fusions with other proteins. This can easily be used as a two hybrid system to detect homo/heterodimers in E. coli, and, in principle, one can use these fusions to turn protein-protein interaction into the repression of transcription. Furthermore, inducible dimerization can lead to a novel switch like behaviors of the system (e.g. [http://2012.igem.org/Team:ETH_Zurich ETH Zurich 2012 team] created a novel UV-B inducible-ON switch).