Tetrameric VP16 transc. activator domain
LexA based yeast light-swithable promoter system
BBa_K801039 (SV40NLS-GAL4AD-Linker-PIF3)
Phytochrome B (PhyB, 908 first N-terminal amino acids)
SV40 NLS (SV40 nuclear localization signal)
BBa_K365005 (20 amino acids long linker)
GAL4AD (GAL4 transcriptonal activation domain)
Fuse lambda cI with LGF2 mutated from Arg to Ala at site 74 (the Glu which locates after three continuous Ala behind lambda repressor is at site 58). As one of the components in Bacterial Two-hybrid System, it causes a stronger interaction with GAL11P compared to the normalized WT.
The LGF2 is mutated from CGA to GCA at site 74.
It is from Agilent Technologies' BacterioMatch ⅡTwo-Hybrid System Vector Kit and modified by Site-Directed Mutagenesis(SDM).
Fuse lambda cI with LGF2 mutated from Asp to Ala at site 78. As one of the components in Bacterial Two-hybrid System, it causes a less interaction with GAL11P compared to the nomalized WT.
From Agilent Technologies' BacterioMatch ⅡTwo-Hybrid System Vector Kit.
GAL4 based yeast light-switchable promoter system
BBa_K801040 (SV40NLS-PhyB-Linker-GAL4DBD)
20 amino acids long linker (BBa_K365005)
SV40NLS-GAL4AD-linker-PIF3 (part for GAL4/LexA based light-switchable promoter system)
CDS for a synthetic hybrid fusion protein. Consists of serveral domains. RFC10 pre and suffix
CDS for a synthetic hybrid fusion protein
SV40 nuclear localization sequence (recognized by importin alpha)
Fuse lambda cI with LGF2 mutated from Asp to Ala at sites 76 and 78 (the Glu which locates after three continuous Ala behind lambda repressor is at site 58). As one of the components in Bacterial Two-hybrid System, it causes little interaction with GAL11P compared to the normalized WT.
The LGF2 is mutated from GAC to GCC at site 76 and from GAC to GCC at site 78.
Fuse lambda cI with LGF2 mutated from Glu to Ala at site 75 (the Glu which locates after three continuous Ala behind lambda repressor is at site 68). As one of the components in Bacterial Two-hybrid System, it causes little interaction with GAL11P compared to the normalized WT.
The LGF2 is mutated from GAA to GCA at site 75.
{| style="color:black" cellpadding="6" cellspacing="1" border="2" align="right" ! colspan="2" style="background:#FFBF00;"|VP64 |- |'''Function''' |transcription activator |- |'''RFC standard''' |[http://parts.igem.org/Help:Assembly_standard_10 RFC 10] |- |'''Backbone''' |pSB1C3 |- |'''Organism''' |''synthetic construct'' |- |'''Source''' |synthetic construct |- |'''Submitted by''' |[http://2016.igem.org/Team:NEU-China NEU-China 2016] |} Tetrameric VP16 transcription activator domain
CooA is a heme-containing transcriptional activator that enables Rhodospirillum rubrum to sense and grow on CO as a sole energy source. CooA is a member of a family of transcriptional regulators similar to the cAMP receptor protein and fumavate nitrate reduction from Escherichia coli. The protein is active in sequence-specific DNA binding in the presence of CO, but not in the absence of CO. The protein to be a dimer in the absence of CO. The product, CooA, is 28% identical (51% similar) to CRP(cAMP receptor protein) and 18% identical (45% similar) to FNR(fumavate nitrate reduction) from Escherichia coli. Inactive Fe(II) CooA structure adapted from that of the strain with PDB identification no. 1FT9. The protein consists of two monomers, shaded differently, which dimerize along the central C-helices of adjacent effector-binding domains. The solved structure is asymmetric, in which one monomer contains fused C- and D-helices. Nonetheless, both F-helices that interact with DNA in a sequence-specific manner are buried from the surface in the structure. The 4/5 loop is noted and so are the Pro2 and His77 heme Fe(II) ligands.
The sequence information was acquired from NCBI and physical DNA was synthesized from GENEART. Classical cloning strategies(restriction digestion) were used to produce biobricks.
Hwan Youn, Robert L. Kerby, Mary Conrad, et al. 2004. Functionally Critical Elements of CooA-Related CO Sensors. J. Bacteriol. 186(5):1320-1329. doi:10.1128/JB.186.5.1320-1329.2004.