Difference between revisions of "Part:BBa K1045001"

Revision as of 17:40, 1 October 2013

DarR ORF with inversed Pre- and Suffix

This is the ORF of DarR (Ms5346). DarR is a transcription factor from Mycobacterium smegmatis, which can bind its specific binding sequence (the DarR operator BBa_K1045000).
The binding effinciency is dependent on the availability of cyclic di-AMP, which enhances the probability of binding to DNA. Ideally, this protein will shut down transcription of information downstream of its operator.
This way, transcription can be controlled via the level of c-di-AMP (e.g. for a reporter-system in order to monitor it).
DarR senses c-di-AMP with its C-terminal domain. DNA-binding is mediated via its N-terminal HTH domain.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NotI site found at 581
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 594
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 31
1000
COMPATIBLE WITH RFC[1000]

@@ Line 2: / Line 2: @@
 <partinfo>BBa_K1045001 short</partinfo>
-This is the ORF of DarR. DarR is a transcription factor from ''Mycobacterium smegmatis'', which can bind its specific binding sequence (the DarR operator [[Part:BBa_K1045000|BBa_K1045000]]). <br/ >
+This is the ORF of DarR (Ms5346). DarR is a transcription factor from ''Mycobacterium smegmatis'', which can bind its specific binding sequence (the DarR operator [[Part:BBa_K1045000|BBa_K1045000]]). <br/ >
 The binding effinciency is dependent on the availability of cyclic di-AMP, which enhances the probability of binding to DNA. Ideally, this protein will shut down transcription of information downstream of its operator. <br/ >
 This way, transcription can be controlled via the level of c-di-AMP (e.g. for a reporter-system in order to monitor it).<br/ >
+DarR senses c-di-AMP with its C-terminal domain. DNA-binding is mediated via its N-terminal HTH domain.
-DarR (Ms5346) was identified in ''Mycobacterium smegmatis''. Since this organism has a high GC content, the coding sequence of DarR is rich in GC, as well. DarR is composed of a C-terminal domain for c-di-AMP-sensing and an N-terminal HTH domain for DNA binding. DarR binds the DarR operator with a footprint of 7 – 8 bp. In EMSA assays, DNA-binding is weak without c-di-AMP. C-di-AMP stimulates DNA binding of DarR.  ''In vitro'', His6–DarR binds c-di-AMP specifically with a Kd = 2.3 ± 0.5 µM. (Zhang ''et al''., 2013)