Difference between revisions of "Part:BBa K1921011"
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===Reference=== | ===Reference=== | ||
[1] Fang S, Pang X, Tian X, et al. BrkAutoDisplay: functional display of multiple exogenous proteins on the surface of Escherichia coli, by using BrkA autotransporter[J]. Microbial Cell Factories, 2014, 14(1):1-12. | [1] Fang S, Pang X, Tian X, et al. BrkAutoDisplay: functional display of multiple exogenous proteins on the surface of Escherichia coli, by using BrkA autotransporter[J]. Microbial Cell Factories, 2014, 14(1):1-12. | ||
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| + | ===Pre-expression=== | ||
| + | <p style="text-align: center;"> | ||
| + | https://static.igem.org/mediawiki/igem.org/1/14/Tjuresults23.jpg<br> | ||
| + | </p> | ||
| + | '''Figure 1.'''This is the pre-expression using E.coli BL21 in different inducing condition.<br> | ||
| + | |||
| + | ===Surface display HPLC results=== | ||
| + | <p style="text-align: center;"> | ||
| + | https://static.igem.org/mediawiki/igem.org/6/60/ProofTJU9.jpg<br> | ||
| + | </p> | ||
| + | '''Figure 2.'''Relative enzyme activity of engineering bacteria E.coli(BL21)/pET22b(+)Brk when induced at 16℃ and 25 ℃ with 0.02mM IPTG.And the last two were induced with 0.09mM IPTG.<br> | ||
Revision as of 05:59, 20 October 2016
BrkA
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 313
Illegal NgoMIV site found at 742
Illegal NgoMIV site found at 1378 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1346
Usage
The anchor of a new autotransportermetthat diated bacterial surface display system BrkAu to Display based on the structure of BrkA. The unique characteristic of BrkA is that the cleaved passenger domain remains adhered on the bacterial surface, which is conducive to keep the displayed exogenous proteins associated onto the cell, thus, the functionalized bacteria can be recycled/re-used conveniently.Now we hope to display our PETase on the surface of E. coli by using it.
Biology
PETase was found from a kind of microorganism(Ideonella sakaiensis 201-F6) living on PET as the main carbon source. It can degrade macromolecular polymers into monomers. PETase is the only enzyme found in bacteria which can degrade PET.
Autotransporter BrkA (Bordetella serum-resistance killing protein A )is from Bordetella pertussis,PETase is the passenger between the signal peptide and the anchor.
Reference
[1] Fang S, Pang X, Tian X, et al. BrkAutoDisplay: functional display of multiple exogenous proteins on the surface of Escherichia coli, by using BrkA autotransporter[J]. Microbial Cell Factories, 2014, 14(1):1-12.
Pre-expression
Figure 1.This is the pre-expression using E.coli BL21 in different inducing condition.
Surface display HPLC results
Figure 2.Relative enzyme activity of engineering bacteria E.coli(BL21)/pET22b(+)Brk when induced at 16℃ and 25 ℃ with 0.02mM IPTG.And the last two were induced with 0.09mM IPTG.