Generator

Part:BBa_K3580102:Design

Designed by: Yuya Otsuki   Group: iGEM20_Waseda   (2020-10-23)


Ptrc-trGPPS-SS(Sabinene synthase)


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 1980
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 1980
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 2366
    Illegal XhoI site found at 4022
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 1980
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 1980
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

This part was designed based on the Ptrc-trGPPS-LS(BBa_K3580101)used in the 2013 paper by Alonso-Gutierrez, Jorge et al. The introduction of the one-amino acid mutation reported in the2015 paper by Srividya, Narayanan et al.The one-amino acid mutation was introduced into the limonene synthase of Mentha spicatato accommodate the N345A mutation, as reported in a 2015 paper by Srividya and Narayanan et al.Actually, limonene synthase used in our project was an E. colicodon-optimized version of a truncated sequence, so a mutation was introduced to change asparagine at position 289 to alanine.

Source

The part BBa_K3580102 is a new part for Parts Registry. It can synthesize sabinene and be used in combination with pBbA5c-MevT-MBI. One amino acid mutation was introduced into limonene synthase of Mentha spicata to create sabinene synthase, by using information reported in a 2015 paper by Srividya and Narayanan et al.

References

(1) Alonso-Gutierrez, J. et al. (2013). Metabolic engineering of Escherichia coli for limonene and perillyl alcohol production. Metabolic engineering, 19, 33-41.

(2) Dudley, Q. M. et al. (2019). Cell-free biosynthesis of limonene using enzyme-enriched Escherichia coli lysates. Synthetic Biology, 4(1), ysz003.

(3) Srividya, N. et al. (2015). Functional analysis of (4S)-limonene synthase mutants reveals determinants of catalytic outcome in a model monoterpene synthase. Proceedings of the National Academy of Sciences, 112(11), 3332-3337.

(4) Korman, T. P. et al. (2017). A synthetic biochemistry platform for cell free production of monoterpenes from glucose. Nature communications, 8(1), 1-8.

(5) Mass Spectrometry Data Center, William E. Wallace, "Mass Spectra" in NIST Chemistry WebBook, NIST Standard Reference Database Number 69, Eds. P.J. Linstrom and W.G. Mallard, National Institute of Standards and Technology, Gaithersburg MD, 20899, https://doi.org/10.18434/T4D303, (retrieved October 22, 2020).