Part:BBa_K3477006

Gene coding for his-tagged PueA enzyme including BBa_0034 RBS, Bba_J23106 promoter and Bba_0010 term

This part constitutes the translational unit of Polyurethanase A (PueA). Upstream of the coding region there is the biobrick prefix, the BBa_J23106 constitutive promoter (from the Anderson family) and the BBa_B0034 ribosome binding site (RBS). The gene sequence comes from Uniprot.org and is codon optimized for Escherichia coli. The sequence is followed by a 6x his-tag sequence. Downstream of the coding region there is the BBa_B0010 terminator and the biobrick suffix.

The enzyme coded for, PueA, originates from Pseudomonas fluorescens ATCC BAA-477 and degrades polyurethane into 4,4-methylenediphenyl-diisocyanate. The his-tag can be used to purify the enzyme using a his-tag purification kit.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal suffix found in sequence at 2030
  Illegal EcoRI site found at 1
  Illegal XbaI site found at 56
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 1
  Illegal NheI site found at 27
  Illegal NheI site found at 50
  Illegal NheI site found at 143
  Illegal SpeI site found at 2031
  Illegal PstI site found at 2045
  Illegal NotI site found at 7
  Illegal NotI site found at 2038
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 1
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal suffix found in sequence at 2031
  Illegal EcoRI site found at 1
  Illegal XbaI site found at 56
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal prefix found in sequence at 1
  Illegal XbaI site found at 56
  Illegal SpeI site found at 2031
  Illegal PstI site found at 2045
  Illegal NgoMIV site found at 1671
  
• 1000
  COMPATIBLE WITH RFC[1000]