Difference between revisions of "Part:BBa K342002:Design"
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+ | "Novel and economical purification of recombinant proteins: Intein-mediated protein purification using in vivo polyhydroxybutyrate (PHB) matrix association", Mahmoud Reza Banki, Tillman U.Gerngross and David W. Wood, 2005, Protein Science, 14:1387–1395 |
Revision as of 16:32, 7 November 2010
Phasin-Phasin Intein (for silver standard fusion)
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Unknown
- 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 1693
- 23INCOMPATIBLE WITH RFC[23]Unknown
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1279
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
This part is a self cleavable fusion protein. It has been designed thanks to the results of the publication
Banki et al.
The phasin protein bind to the granule membrane. The second phasin makes the binding stronger and then increases the rate of purified protein during the step of granules extraction. The intein is a self cleavable protein which breaks under PH=6 and allows the release od the fused protein at the end of the sequence The prefix biobrick standard used contains a perfect RBS. This part should had been synthesized by Mr gene in once but due to technical issues we had to redesign two separates fusion biobrick and ligate them. The sequencing of this part confirm the expected sequence.
The suffix is a silver standard fusion suffix : 5' ACTAGT A GCGGCCG CTGCAG 3'.
The phasin sequence is the same as the biobrick BBa_K208001 from the Utah 2009 team.
The intein is from the self cleaving protein sequence delta.I-CM mini intein from mycobacterium tuberculosis described in this patent .
The in between are the same as the one described in Banki et al.
Source
Ligation of two synthetized parts : Phasin and Phasin-Intein
References
"Novel and economical purification of recombinant proteins: Intein-mediated protein purification using in vivo polyhydroxybutyrate (PHB) matrix association", Mahmoud Reza Banki, Tillman U.Gerngross and David W. Wood, 2005, Protein Science, 14:1387–1395