Difference between revisions of "Part:BBa K2332002"

Revision as of 17:19, 24 October 2017

Blue light inducible promoter (Pblind)

Promoter activated by EL222 (BBa_K2332004) upon blue-light exposure (450nm)

Pblind is a blue-light inducible promoter that will only allow RNAP to transcribe genes downstream upon blue-light (450) exposure, as this will induce a conformational change in EL222, a natural photosensitive DNA-binding protein, required for RNAP recruitment and transcriptional activation. To enable blue-light dependent transcriptional control, EL222 should be constitutively expressed by the cells.

Since light can be controlled easily in space, time and degree, this new basic part will enable tight spatiotemporal control of gene expression.

Usage and Biology

This part consists of a fusion of the EL222 DNA binding region and the LuxI promoter. The lux box, a 20bp inverted repeat (LuxR and 3-oxo-C6-HSL complex binding region) from the luxI promoter, was replaced with the 18bp DNA binding region of EL222. In the dark, EL222 is inactive as its N-terminal LOV domain represses its DNA-binding C-terminal HTH domain. Upon blue light exposure (450nm), LOV-HTH interaction is released, allowing it to dimerize and bind its binding region, overlapping the -35 region of the luxI promoter. This ultimately results in the recruitment of RNAP and transcriptional activation.

Figure 1: Blue light inducible expression system. Under blue light, the EL222 DNA binding protein dimerises and bind its binding region within the designed Pblind promoter, overlapping the -35 region of the luxI promoter. This ultimately results in the recruitment of RNAP and transcriptional activation. Figure adapted from Jayaraman P. et al. (2016)

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

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 <partinfo>BBa_K2332002 short</partinfo>
-Pblind is a blue light inducible promoter. It requires cells to also express EL222 ([https://parts.igem.org/Part:BBa_K2332004 BBa_K2332004]), a natural photosensitive DNA-binding protein that becomes active only upon blue-light exposure. This part consists of a fusion of the EL222 DNA binding region and the LuxI promoter. The lux box, a 20bp inverted repeat (LuxR and 3-oxo-C6-HSL complex binding region) from the luxI promoter, was replaced with the 18bp DNA binding region of EL222. In the dark, EL222 is inactive as its N-terminal LOV domain represses its DNA-binding C-terminal HTH domain. Upon blue light exposure (450nm), LOV-HTH interaction is released, allowing it to dimerize and bind its binding region, overlapping the -35 region of the luxI promoter. This ultimately results in the recruitment of RNAP and transcriptional activation.
+Promoter activated by EL222 ([https://parts.igem.org/Part:BBa_K2332004 BBa_K2332004]) upon blue-light exposure (450nm)
+Pblind is a blue-light inducible promoter that will only allow RNAP to transcribe genes downstream upon blue-light (450) exposure, as this will induce a conformational change in  EL222, a natural photosensitive DNA-binding protein, required for RNAP recruitment and transcriptional activation. To enable blue-light dependent transcriptional control, EL222 should be constitutively expressed by the cells.
+Since light can be controlled easily in space, time and degree, this new basic part will enable tight spatiotemporal control of gene expression.
 ===Usage and Biology===
+This part consists of a fusion of the EL222 DNA binding region and the LuxI promoter. The lux box, a 20bp inverted repeat (LuxR and 3-oxo-C6-HSL complex binding region) from the luxI promoter, was replaced with the 18bp DNA binding region of EL222. In the dark, EL222 is inactive as its N-terminal LOV domain represses its DNA-binding C-terminal HTH domain. Upon blue light exposure (450nm), LOV-HTH interaction is released, allowing it to dimerize and bind its binding region, overlapping the -35 region of the luxI promoter. This ultimately results in the recruitment of RNAP and transcriptional activation.
 [[File:Pblind-GFP.png|thumb|center|500px| Figure 1: Blue light inducible expression system. Under blue light, the EL222 DNA binding protein dimerises and bind its binding region within the designed Pblind promoter, overlapping the -35 region of the luxI promoter. This ultimately results in the recruitment of RNAP and transcriptional activation. Figure adapted from [https://www.ncbi.nlm.nih.gov/pubmed/27353329 Jayaraman P. et al. (2016)]]]