Composite
rAIP

Part:BBa_K2027000:Design

Designed by: Charles Gleason, Michael Becich   Group: iGEM16_Stanford-Brown   (2016-09-07)


Recombinant Apoptosis-Inducing Protein (L-lysine Alpha Oxidase)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 1004
    Illegal BglII site found at 1032
    Illegal BamHI site found at 767
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 25


Design Notes

As far as design goes, this is mostly just a native Scomber japonicus enzyme codon-optimized for Escherichia coli. The conjugated tag should allow for purification and visualization with anti-FLAG antibodies, visualization with Lumio™ Green, and purification with nickel columns. Tani et al. characterized this enzyme in their quest for synthesis of L-pipecolic acid from racemic lysine after nickel column purification,1 and we verified the function of our part in a similar way. More information can be found on the experience page for this part. Below is the forward sequencing verification for the part; enzyme functionality and successful purification indicate that the carboxy-terminus is also intact.

T--Stanford-Brown--BBa_K2027000-Sequence-Verification.png

Source

Tani, Y., Miyake, R., Yukami, R. et al. Appl Microbiol Biotechnol (2015) 99: 5045. doi:10.1007/s00253-014-6308-0

References

1. Tani, Y., Miyake, R., Yukami, R. et al. Appl Microbiol Biotechnol (2015) 99: 5045. doi:10.1007/s00253-014-6308-0