Part:BBa_K4444011
PhoCl Display mRuby
PhoCl Display mRuby Backbone is modified from the RELEASE system. It contains a mRuby as the protein of interest, PhoCl2c, a tri-transmembrane domain and an ER retention motif. The furin cut site, HCV cut site, and GFP were removed. PhoCl Display will display a protein of interest on the cell membrane in response to ultraviolet light stimulation.
Usage and Biology
Our project builds on the protein circuit RELEASE (Vlahos et al.) by introducing light-inducible proteins or photoswitches. Photocleavable protein (PhoCl) is a recently developed optogenetic technology that self-cleaves when stimulated by 405 nm ultraviolet light. PhoCl was engineered from a green-to-red photoconvertible fluorescent protein. When stimulated with ultraviolet light, the protein undergoes a β-elimination reaction, producing a small peptide fragment and a large barrel fragment that will spontaneously dissociate (Zhang et al.).
Mechanism: Upon light-activation Phocl breaks, removing the ER retention motif and allowing the protein to continue along the secretion pathway. The absence of the furin cut site means the POI remains bound to the membrane of the vesicle. At the plasma membrane the POI will be displayed.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 52
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 772
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 895
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1417
Illegal SapI.rc site found at 73
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