Part:BBa_K1129046

4-coumarate 3-hydroxylase under pTET constitutive promoter

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 100
  Illegal NgoMIV site found at 108
  Illegal NgoMIV site found at 858
  Illegal NgoMIV site found at 1031
  Illegal NgoMIV site found at 1433
  Illegal NgoMIV site found at 1437
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 632
  

Background

Ferulic acid is best utilized as a precursor for synthesis of aromatic compounds such as vanillin, which involves 2 enzymes: the conversion of ferulic acid to feruloyl-CoA by feruloyl-CoA synthetase (Fcs) enzymes and the conversion of feruloyl-CoA into vanillin by enoyl-CoA hydratase/aldolase (Ech) enzymes BBa_K1129002 [1]. BBa_K1129001 is a feruloyl-CoA-synthetase (fcs) from Pseudomonas putida KT2440 which converts ferrulic acid to feruloyl CoA [2].

Experimental Data

4-coumarate 3-hydroxylase is an enzyme in the vanillin pathway. 4-coumarate 3-hydroxylase (4-CMH) hydrolyses p-coumaric acid to caffeic acid. It is the 2nd enyzme the biosynthetic pathway of vanillin from the amino acid tyrosine.

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Figure 3. Compound generation identification by GC-MS. Chromatograms and mass spectra for select peaks are shown. Structures represent predictions based on library matching or comparison to standards. Controls represent plasmids missing the gene of interest. A) Internal control using caffeic acid. B) Conversion of p-coumaric acid to caffeic acid by constitutive expressed 4CMH. C) Possible conversion of p-coumaric acid to caffeic acid by inducibly expressed 4CMH. The mass spectrum however is confounded by another compound.