Coding

Part:BBa_K4722020

Designed by: Anqi Chen   Group: iGEM23_BJEA-China   (2023-10-09)


SP-Histag-NicX-BrkA

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 971
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 971
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 971
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 971
    Illegal NgoMIV site found at 1130
    Illegal NgoMIV site found at 1823
    Illegal NgoMIV site found at 2252
    Illegal NgoMIV site found at 2888
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 2856

Usage and Biology

SP[1][2], Histag, NicX[3][4], BrkA[5] were fused together

Design Consideration

The construction includes: NicX with SP, BrkA and Histag NicX was genetically connected with SP and BrkA to enable its direct translation onto the surface of BL21. This innovation eliminated the need for protein purification steps, allowing for the direct utilization of E. coli as a host for enzymes in various applications. Enzymatic cleavage was performed at the NcoI and XhoI restriction sites, allowing for the precise integration of SP-Histag-NicX-BrkA. The part contains Histag and does not require Histag on the plasmid.

References

  1. Wang, S. N., Liu, Z., Tang, H. Z., Meng, J., & Xu, P. (2007). Characterization of environmentally friendly nicotine degradation by Pseudomonas putida biotype A strain S16. Microbiology, 153(5), 1556-1565. https://doi.org/10.1099/mic.0.2006/005223-0
  2. Wang, W., Xu, P., & Tang, H. (2015). Sustainable production of valuable compound 3-succinoyl-pyridine by genetically engineering Pseudomonas putida using the tobacco waste. Scientific Reports, 5(1), 16411. https://doi.org/10.1038/srep16411
  3. Chen, B., Sun, L., Zeng, G., Shen, Z., Wang, K., Yin, L., ... & Jiang, C. (2022). Gut bacteria alleviate smoking-related NASH by degrading gut nicotine. Nature, 610(7932), 562-568. https://doi.org/10.1038/s41586-022-05299-4
  4. Jiménez, J. I., Canales, Á., Jiménez-Barbero, J., Ginalski, K., Rychlewski, L., García, J. L., & Díaz, E. (2008). Deciphering the genetic determinants for aerobic nicotinic acid degradation: the nic cluster from Pseudomonas putida KT2440. Proceedings of the National Academy of Sciences, 105(32), 11329-11334.https://doi.org/10.1073/pnas.080227310
  5. Sun, F., Pang, X., Xie, T., Zhai, Y., Wang, G., & Sun, F. (2015). BrkAutoDisplay: functional display of multiple exogenous proteins on the surface of Escherichia coli by using BrkA autotransporter. Microbial Cell Factories, 14, 1-12. https://doi.org/10.1186/s12934-015-0316-3
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