Part:BBa_K2671420
TetR-TetP-GroES
The gene coding for the chaperone GroES (homologue to HSP10 in eukaryotes) that is found in E.coli. It often works in conjunction to GroEL creating an environment well suited for the folding of proteins. NOTE: The sequence is codon optimized (E.coli) with IDT's tool. This sequence will not yield any results with BLASTn, but will hit GroES with BLASTx. The TetP also has one TetR binding site mutated (one base).
The gene coding for GroES is placed downstream from a tetracycline promotor. Further upstream the tetracycline repressor protein (TetR) is found, expressed by a constitutive promotor. The TetR protein has a termination sequence directly downstream from it, while the GroES gene has its transcription stop by the E.coli his operon termination sequence that is present in most pSB vectors, directly downstream of the insert.
Usage and Biology
The results for our engineered system is shown below. Our biobrick is only called GroES in the graphs for simplicity. The substrate proteins has their names written out. GroE is a plasmid where both GroEL and GroES is expressed. The concentrations we used to induce all the different systems was: 200 ng/ml tetracycline for BBa_K2671420 (GroES), 0.5 mg/ml L-arabinose for the GroE plasmid and 0.5 mM IPTG for all the different substrates. All measurements was done in vivo in a 96-well plate. Excitation was done at 485 nm for all substrates and emission was measured at 520 nm. The chaperone plasmids, this biobrick and GroE was induced 30 mins prior to the substrates. Absorbance at 600 nm was measured once in the start and at the end of the 16 hour experimental time.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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