Part:BBa_K3887003:Design

PyrH

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NotI site found at 1404
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 211
  Illegal NgoMIV site found at 829
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1384
  Illegal BsaI.rc site found at 1354
  Illegal SapI site found at 426
  

Design Notes

PyrH is another type of halogenase enzyme used also during conversion of indole. In our project, it is used to biosynthesis other types of indigo using Trp 7-halogenase, similar to SttH producing Tyrian Purple. It is applied with the Fre-L3-halogenasee fusion technology. The Fre-L3-PyrH is placed in the TnaA action to convert to indole. Then, it is applied into the two-cell reaction system, with 2.5mM Trp as initiate substrate.

Based on experiment, 7,7′-dichloroindigo (0.26 mM, 9), and 7,7′-dibromoindigo (0.13mM, 10) were produced from Trp 7-halogenase pyrH and Fre-L3-PyrH. As a result, several colors of halogenated indigos could be produced.

References

1.2.Phillips, R. S., Buisman, A. A., Choi, S., Hussaini, A., & Wood, Z. A. (2018). The crystal structure of Proteus vulgaris tryptophan indole‐lyase complexed with oxindolyl‐l‐alanine: implications for the reaction mechanism. Acta Crystallographica. Section D, Structural Biology, 74(8), 748–759. 2.Lee, J., Kim, J., Song, J.E. et al. Production of Tyrian purple indigoid dye from tryptophan in Escherichia coli. Nat Chem Biol 17, 104–112 (2021).