Team:Penn/SurfaceDisplayBBa

Ice nucleation protein (INP) is a protein found in Xanthomonas campestris pc. campestris BCRC 12846. Its function is to provide a surface for ice nucleation, which results in the formation of ice crystals. However, recent studies have utilized INP for its surface display properties. In nature, the protein is anchored in the membrane through a glycosylphosphatidylinositol (GPI) anchor, a relatively rare occurrence in prokaryotes.

Part BBa_K811005 has been utilized by the 2012 Penn iGEM team to display a variety of different proteins. The red fluorescent protein mCherry has been successfully displayed on the surface of E. Coli, where it can produce fluorescence. We displayed mCherry on the outer membrane of E. coli BL21. After sonication and centrifugation of induced cells, almost all of the mCherry was localized in the membrane fraction when fused to INPNC, whereas in the control Intein-mCherry fusion (which exhibits cytoplasmic localization), all of the mCherry was contained in the lysate (Figure 1).

Figure 1

Figure 1: Surface display of mCherry using INPNC system. INPNC-mCherry and Intein-mCherry fusions were expressed in E. coli BL21 in the pET26b expression vector and Wood-Intein expression plasmid, respectively. When fused to INPNC, almost all mCherry was localized in the membrane fraction after sonication and centrifugation, while in the case of Intein-mCherry, all mCherry was localized in the cytoplasmic lysate.