Difference between revisions of "Part:BBa K1075024"

Latest revision as of 01:26, 5 October 2013

RBS34- mCherry- (Ec)ssrA(DAS+4)-TT

The part contains a ribosomal binding site, the red fluorescent protein mCherry fused to the E. coli(ec)ssrA (DAS+4) tag and a a double terminator.

biology

The (ec)ssrA tag is a short peptide sequence that can be fused to the C-terminus of proteins which should be degraded. The adaptor protein SspB binds to it and delivers the substrates to the protease ClpXP complex in E. coli. The mutation (DAS+4) in (ec)ssrA prevents the degradation of the ssrA-tagged protein without the presence of sspB and therefore makes the system more specific and controllable. [http://www.ncbi.nlm.nih.gov/pubmed/16762842]

mCherry is a red fluorescent protein with the excitation maximum at 587 nm and the Emission maximum at 610 nm. [http://www.ncbi.nlm.nih.gov/pubmed/15558047]

application

As we want to control protein degradation by controlling the function of ecSspB, we tagged the red fluorescent protein mCherry with ecssrA(DAS+4) to measure the degradation rate.

The part was designed for proof of principle. Application as a bacterial fotographic film might be possible as well.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 1509
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

@@ Line 2: / Line 2: @@
 <partinfo>BBa_K1075024 short</partinfo>
-RBS34-mCherry-ecssrA(DAS+4)-TT
+The part contains a ribosomal binding site, the red fluorescent protein mCherry fused to the E. coli(ec)ssrA (DAS+4) tag and a a double terminator.
-===construction===
-The part contains a ribosomal binding site, the red fluorescent protein mCherry fused to the ssrA (DAS+4) tag and a double terminator.
 ===biology===
-The ribosomal binding side is a sequence of mRNA where the ribosome binds to start translation.
+The (ec)ssrA tag is a short peptide sequence that can be fused to the C-terminus of proteins which should be degraded. The adaptor protein SspB binds to it and delivers the substrates to the protease ClpXP complex in E. coli. The mutation (DAS+4) in (ec)ssrA prevents the degradation of the ssrA-tagged protein without the presence of sspB and therefore makes the system more specific and controllable. [http://www.ncbi.nlm.nih.gov/pubmed/16762842]
-The ssrA tag is a short peptide sequence, which is fused to the C-terminus of proteins, which should be degraded. It relates to the protease ClpXP complex in E.coli and it also allows the adaptor proteins SspB binding and delivering substrates to the proteases in order to make the process more efficient. The mutated ssrA (DAS+4) weakens a direct binding between proteases and ssrA and increases the dependance of sspB. [http://www.ncbi.nlm.nih.gov/pubmed/16762842]
 mCherry is a red fluorescent protein with the excitation maximum at 587 nm and the Emission maximum  at 610 nm. [http://www.ncbi.nlm.nih.gov/pubmed/15558047]
-The double terminator stops the transcription at this point.
 ===application===
-As we want to control protein degradation by controlling the function of SspB, we tag the red fluorescent protein mCherry with ssrA (DAS+4) to measure the degradation rate.
+As we want to control protein degradation by controlling the function of ecSspB, we tagged the red fluorescent protein mCherry with ecssrA(DAS+4) to measure the degradation rate.
-Application as a bacterial fotographic film might be possible as well.
+The part was designed for proof of principle. Application as a bacterial fotographic film might be possible as well.
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