Part:BBa_K1357006

Manganese Peroxidase Expression Vector

Manganese peroxidase (MnP) is an enzyme from the fungus, Phanerochaete chrysosporium. This enzyme has been shown to degrade nylon 6 and nylon 66 (nylon 66 more effectively). MnP helps lignin degradation in Phanerochaete chrysosporium. The sequence for MnP in this BioBrick has been codon optimized for E.coli. A pelB secretion tag was also attached to the MnP sequence. The BioBrick also contains a constitutive promoter, a ribosome binding site and a terminator. This BioBrick allows E.coli to secrete manganese peroxidase into the surrounding environment. It is intended for use by teams that are exploring plastic degradation in their research. Enzymatic activity requires a pH near 4.5 as well as a manganese source such as manganese sulphate. Works best at temperatures near 32 C. Related genes include nylC, nylon hydrolaze, which is also used to degrade nylon plastics.

The chart below shows that MnP has been secreted out of the cell. As it is excreted out of the cell it oxidizes Syringaldazine producing a red color and increasing OD readings at 530nm

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These images illustrates the effect MnP has on nylon-6,6 after 1 day of incubation. The images on the left are nylon-6,6 incubated with control BL-21 cells with pSB1C3 transformed into them. The images on the left are treated with BL-21 cells that have been transformed with K1357006. It was incubated for 24 hours at 32℃ at 160 rpm with a pH of 4.5. While the differences look minimal at lower magnifications, at higher magnifcations we see a high degree of aberration indicating that the nylon is in fact being broken down.

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 116
  Illegal NgoMIV site found at 1034
  Illegal AgeI site found at 783
  Illegal AgeI site found at 828
  Illegal AgeI site found at 1026
  Illegal AgeI site found at 1056
  
• 1000
  COMPATIBLE WITH RFC[1000]