Team:EPFL/Improve

iGEM EPFL 2018

Improve

Introduction

Encapsulins are versatile proteins found in a variety of different bacteria (Giessen and Silver, 2017). In the case of this specific part derived from Thermotoga maritima, it can be used among other things to deliver cargo, both on the outer surface of the cage by fusing a peptide in between the 139/140 Amino Acids or the protein's C terminus. A cargo protein can also be loaded by fusing it with a tag binding to Encapsulin's interior surface (Cassidy-Amstutz et al., 2016).

BBa_K2696005

BBa_K192000 is a part that was brought to the registry by iGEM Toronto 2009. We designed an upgraded composite part, BBa_K2686005 using a modified version of David Savage's encapsulin (Addgene #86405) where we introduced a HexaHistidine loop on the protein subunit's inner surface which increases the heat resistance and stability of the part. To facilitate the introduction of antigen encoding sequences into the HexaHistidine Encapsulin, BsaI cut sites are present on the Ecapsulin's C terminus with an insert coding for sfGFP under a native promoter to enable selection of successful transformants by color.

Bibliography

Cassidy-Amstutz, C., Oltrogge, L., Going, C., Lee, A., Teng, P., Quintanilla, D., East-Seletsky, A., Williams, E. and Savage, D. (2016). Identification of a Minimal Peptide Tag for in Vivo and in Vitro Loading of Encapsulin. Biochemistry, 55(24), pp.3461-3468. Giessen, T. and Silver, P. (2017). Widespread distribution of encapsulin nanocompartments reveals functional diversity. Nature Microbiology, 2, p.17029.