Coding

Part:BBa_K1499201

Designed by: Raman Nelakanti   Group: iGEM14_StanfordBrownSpelman   (2014-08-25)

Deinococcus radiodurans uracil-DNA glycosylase 1

This part encodes a uracil-DNA glycosylase found in D. radiodurans, which protects cells from radiation-induced DNA damage. In the literature, it is referred to as DR0689 and detailed protein features can be found on its NCBI Protein page.

Usage and Biology

Deinococcus radiodurans is a highly radiation-resistant bacterium that can survive after up to 12,000 Gy (absorbed radiation dose, Gray) of radiation. 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. D. radiodurans is known to survive both ionizing and UV radiation.

One gene that contributes to this phenotype is a family 1 uracil-DNA glycosylase (UDG), hereafter called uracil-DNA glycosylase 1 (UDG1), which catalyzes the excision and repair of cytosine->uracil mutations that occur following UV exposure (1)(2). The enzyme can remove uracil from U:G and U:A pairs in both double-stranded and single-stranded DNA. Out of the four families of UDGs, family 1 UDGs are found in E. coli, humans, and DNA viruses (1).


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 720
  • 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 (Figure 1).

Figure 1. UDG1 sequence matches the expected sequence from the genomic DNA.

Results

Functional data are in the process of being obtained. Transformation of the part ligated to a constitutive promoter and medium RBS failed twice, suggesting that it may be toxic if it is highly expressed.


References

1. Sandigursky, M et al. (2004) Multiple uracil-DNA glycosylase activities in Deinococcus radiodurans. DNA Repair (Amst) 3(2):163-9. PMID: 14706350.
2. Pearl, LH (2000) Structure and function in the uracil-DNA glycosylase superfamily. Mutat. Res. 460(3-4):165-81. PMID: 10946227.

[edit]
Categories
//cds
//chassis/prokaryote/ecoli
Parameters
biologyDeinococcus radiodurans
proteinuracil-DNA glycosylase