Part:BBa_K1976052
Colicin E2 DNase domain and immunity protein Im2 generator with anderson promoters
Generator of colicin E2 DNase domain BBa_K1976048 and immunity protein Im2 BBa_K1976026.
Figure 1: The structure of the Im2 immunity protein and Colicin E2 in a complex.
Usage
This part can be used to produce colicin without killing the host cell, as the immunity protein is produced more than equimolar to the DNase fragment. If BBa_K1976028 is used instead of BBa_K1976027, it can function as a amber-suppression inducible killswitch.
Characteristics
Figure 2: SDS-PAGE of the Colicin E2 DNase domain, the blue arrow marks the band of the expression. From left to right: Top10 E. coli, Top10 E. coli transformed with BBa_K1976052 and Top10 E. coli transformed with BBa_K1976053.
Figure 3: 1% agarose gel electrophoresis, stained with HD-green. The protein product from this part was cleaned up and used in different concentrations to test its activity. From left to right: pSB1C3 alone, pSB1C3 in the lysate buffer 1x, pSB1C3 in the lysate buffer 3x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976052 1x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976052 3x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976053 1x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976053 3x.
| Molecular Weight |
Colicin E2 Dnase domain
15.2 kDa Immunity Protein 10.0 kDa |
| Residues | Colicin E2 Dnase domain 133 |
| pI |
Colicin E2 Dnase domain
9.06 Immunity Protein 4.66 |
| [Data taken created in Snapgene and http://isoelectric.ovh.org/calculate.php] |
Characteristics
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30
Illegal NheI site found at 477
Illegal NheI site found at 500 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 681
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 89
- 1000COMPATIBLE WITH RFC[1000]
[1] Cascales et al, Colicin Biology, Microbiology and Molecular Biology Reviews, vol. 71, pp. 158-229, 2007
| None |