Part:BBa_K4385015
Copper constitutive surface display
This part was mainly composed of constitutive promoter J23100, the anchoring protein OmpC-CBP and the heavy metal detoxification protein. Promoter J23100 continuously expressed downstream copper ion surface display proteins to verify the function of outer membrane protein C and lipoprotein of copper ion surface display system. The 6*His Tags were added to this sequence to purify the recombinant protein.
Usage and Biology
Metallothioneins (MTs) are a family of heavy metal detoxification protein. To counter the detrimental effect of heavy metal on our engineered bacteria, MTs will be continuously expressed in our engineered bacteria to increase their tolerance to heavy metals.Promoter J23100 continuously expressed downstream copper ion surface display proteins to verify the function of outer membrane protein C and lipoprotein of copper ion surface display system. The 6*His Tags were added to this sequence to purify the recombinant protein.
Characterization
To verify the effect of soluble MTs have on the cells’ metal tolerance abilities, two plasmids, PJ23100-OmpC-CBP and PJ23100-OmpC-CBP-PJ23100-ST,which was shorted for PJ23100-OmpC-CBP-ST, were constructed. E. coli harboring PJ23100-OmpC-CBP-ST and PJ23100-OmpC-CBP were subjected to medium containing different concentrations of copper ions. OD600 measurements were conducted to analyze the effects of ST on bacterial growth. As shown in Fig. 1C, ST enhanced the tolerance of bacteria in heavy metal solutions. Therefore, we added ST in all other composite parts of surface display system.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 414
Illegal SpeI site found at 573
Illegal PstI site found at 921 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 414
Illegal NheI site found at 7
Illegal NheI site found at 30
Illegal NheI site found at 1289
Illegal NheI site found at 1312
Illegal SpeI site found at 573
Illegal PstI site found at 921 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 414
Illegal BglII site found at 122
Illegal BglII site found at 1424 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 414
Illegal SpeI site found at 573
Illegal PstI site found at 921 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 414
Illegal SpeI site found at 573
Illegal PstI site found at 921
Illegal AgeI site found at 240 - 1000COMPATIBLE WITH RFC[1000]
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