Difference between revisions of "Part:BBa K2686000"
Line 17: Line 17:
 
<partinfo>BBa_K2686000 parameters</partinfo>
 
<partinfo>BBa_K2686000 parameters</partinfo>
 
<!-- -->
 
<!-- -->
 +
 +
==Characterization==
 +
 +
A variety of different characterization techniques were used to assess the properties of the encapsulin protein cage.
 +
 +
===Assembly===
 +
 +
The self assembly of the encapsulin 60-mer was first examined using SDS PAGE, where a high band is expected to form due to the high molecular weight and size of the 1.3 MDalton complex.
 +
Multiple gels were made, some containing the denatured cell free reaction mixture, others containing the same but after a variety of purification procedures.
 +
 
===References===
 
===References===
 
Choi, B., Moon, H., Hong, S., Shin, C., Do, Y., Ryu, S. and Kang, S. (2016). Effective Delivery of Antigen–Encapsulin Nanoparticle Fusions to Dendritic Cells Leads to Antigen-Specific Cytotoxic T Cell Activation and Tumor Rejection. ACS Nano, 10(8), pp.7339-7350.
 
Choi, B., Moon, H., Hong, S., Shin, C., Do, Y., Ryu, S. and Kang, S. (2016). Effective Delivery of Antigen–Encapsulin Nanoparticle Fusions to Dendritic Cells Leads to Antigen-Specific Cytotoxic T Cell Activation and Tumor Rejection. ACS Nano, 10(8), pp.7339-7350.
  
 
Moon, H., Lee, J., Min, J. and Kang, S. (2014). Developing Genetically Engineered Encapsulin Protein Cage Nanoparticles as a Targeted Delivery Nanoplatform. Biomacromolecules, 15(10), pp.3794-3801.
 
Moon, H., Lee, J., Min, J. and Kang, S. (2014). Developing Genetically Engineered Encapsulin Protein Cage Nanoparticles as a Targeted Delivery Nanoplatform. Biomacromolecules, 15(10), pp.3794-3801.

Revision as of 16:20, 13 October 2018


Encapsulin with HexaHistidine insert and C-terminal OT1

This part codes a modified Thermotoga maritima Encapsulin protein. The part is optimized for expression in E. coli and has an additional HexaHistidine insert between amino acids 43 and 44, forming a loop on the interior surface of the encapsulin monomer providing higher heat resistance and stability, and better hydrodynamic properties (Moon et al., 2014). The C-terminus of the encapsulin is fused to a SIINFEKL (OVA) peptide which is displayed on the exterior surface of the encapsulin momomer as an antigen (Choi et al., 2016).

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 77
    Illegal BglII site found at 492
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 426
    Illegal SapI.rc site found at 457


Characterization

A variety of different characterization techniques were used to assess the properties of the encapsulin protein cage.

Assembly

The self assembly of the encapsulin 60-mer was first examined using SDS PAGE, where a high band is expected to form due to the high molecular weight and size of the 1.3 MDalton complex. Multiple gels were made, some containing the denatured cell free reaction mixture, others containing the same but after a variety of purification procedures.

References

Choi, B., Moon, H., Hong, S., Shin, C., Do, Y., Ryu, S. and Kang, S. (2016). Effective Delivery of Antigen–Encapsulin Nanoparticle Fusions to Dendritic Cells Leads to Antigen-Specific Cytotoxic T Cell Activation and Tumor Rejection. ACS Nano, 10(8), pp.7339-7350.

Moon, H., Lee, J., Min, J. and Kang, S. (2014). Developing Genetically Engineered Encapsulin Protein Cage Nanoparticles as a Targeted Delivery Nanoplatform. Biomacromolecules, 15(10), pp.3794-3801.