Part:BBa_K4821003

RdhANp_M11_K12

This sequence codes for a modified RdhANp reductive dehalogenase that has been docked to PCB-11 and codon-optimized for expression in E. coli K-12. This part contains the following mutations: K295L K310A R312T P314A M315L S423T S425V M426L N455L H457D R555M V556I A563G. It was designed as an improvement on part BBa_K4821002, the codon-optimized wild-type RdhANp dehalogenase to possess high catalytic activity against PCB11.

Usage and Biology

This is a modified RdhANp with the potential to dechlorinate PCB11 for more efficient breakdown of the compound to acetyl CoA by the biphenyl degradation pathway (bph gene cluster). Mechanistically, Y426 in RdhANp protonates the chlorine located at 3.5A from the oxygen of Y426 and 5.0A from the cobalamin to catalyze a dechlorination reaction where the chlorine would coordinate with the Co on the cobalamin.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 1628
  Illegal BamHI site found at 312
  Illegal BamHI site found at 419
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]