Part:BBa_K2271105:Experience

Import of fluorescent proteins into the peroxisomal matrix

Figure 1:PEX5 knock out strain, transformed with the R15 variant, mTurqouise tagged with the PTS1 and the PEX13 membrane anchor fused to mRuby.

Figure 2:PEX5 knock out strain, transformed with the R15 variant, mTurqouise tagged with the PTS1* P3 and the PEX13 membrane anchor fused to mRuby.

The figures 1 and 2 show two different experiments. The first in figure 1 is the control if the PEX5 variant R15 imports proteins tagged with the natural PTS1 and the second in figure 2 is an actual experiment if the variant imports proteins tagged with one of our new PTS1* variants − in this case it was P3 (-YTNQE). One can see that there is no import in both cases. While PEX13-mRuby marks the peroxisomal membrane, mTurqouise is located in the cytosol − this indicates that there is no import.
We wanted to make a screening for functional PTS1* variants but unfortuneately the Violacein assay planned for that (see our [http://2017.igem.org/Team:Cologne-Duesseldorf/Design#h3-3 wiki article] for more information) did not work out. As outlook we propose to find a fitting PTS1* hence the molecular dynamics simulations showed promising results.

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