Part:BBa_K1499205

MntH with amber stop codon with promoter for expression, no tRNA

This part encodes the manganese transporter, MntH, with one UAG stop codon in place of a leucine and a promoter for expression. MntH is involved with manganese homeostasis, a critical process for dealing with free radicals generated upon exposure to radiation that could potentially cause DNA damage.

Usage and Biology

Deinococcus radiodurans is a highly radiation-resistant bacterium that can survive after up to 12,000 Gy (absorbed radiation dose, Gray). The unique genes that allow it to better handle radiation exposure may provide ways to make E. coli and other organisms more resistant for applications of synthetic biology in tough environmental conditions, like space. Manganese is a cofactor for many of the radiation stress mechanisms in the cell, and so manganese transporters bring manganese into the cell during radiation stress. These transporters are particularly well-known in D. radiodurans, and expressing these proteins in E. coli has shown promise [1].

We have observed that this coding part imparts radiation resistance when expressed under a strong constitutive promoter and medium RBS. Although we have not tried inducible or weaker promoters, we expect the part to work under these conditions.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 215
  Illegal NgoMIV site found at 1007
  Illegal NgoMIV site found at 1080
  
• 1000
  COMPATIBLE WITH RFC[1000]

Characterization

Verification of Part

The part was sequence verified in the pSB1C3 backbone before submission to the registry. Two reads, forward and reverse, were obtained using VF2 and VR.The site-directed mutagenesis worked at our targeted TGA->TAG stop codon mutation.

Results

The part has been characterized.

References

1. Haiyan, S & Baoming, T (2010). Radioresistance analysis of Deinococcus radiodurans gene DR1709 in Escherichia coli.African Journal of Microbiol. Research, 4(13): 1412-1418.