Coding

Part:BBa_K4586005

Designed by: Mohamed Mohamed Saad Aboelghare   Group: iGEM23_AFCM-Egypt   (2023-09-15)
Revision as of 05:07, 25 September 2023 by Ahmed Mattar (Talk | contribs)


MCP-ADAR2

Part Description

This part codes for the catalytic portion of ADAR2 (adenosine deaminases acting on double-stranded RNA) that converts adenosines (A) to inosines (I) in both coding and non-coding RNA transcripts. Through a process known as hybridization or deamination, In addition to conjugating this part to MCP (MS2 coat protein), which has an affinity for the MS2 hairpin structure.

Usage

This part is implemented in our design to increase the effectivity and sensitivity of our therapeutic agent, as changing the status of the DART V ADAR switch from off to on is mediated through the ADAR enzyme by deamination of the mismatched adenosine group into an inosine group. Thus, the stop Codon UAG within the sensor of the switch will be converted into UIG, and the translation of the therapeutic agent will precede expressing Cas12k and MCP-ADAR2. Thus, MCP-ADAR2 is creating a positive feedback loop that amplifies the signal transmitted from different cargo copies present within the target cell as shown in figure 1.

Figure 1: This figure illustrate the activity of our DART V ADAR tissue specific switch that is designed to be in the on state after recognition of the autoreactive B-cells,this recognition based on mismatched base editing in the level of transcribed RNA that is mediated through ADAR enzyme activity.

Literature Characterization

The study tested the action of sensors containing MS2 hairpins without ADAR, with ADAR p150, or with MCP-ADAR2dd.

Off-state refers to mNeonGreen expression in the absence of iRFP720 trigger mRNA, while on-state refers to mNeonGreen expression in the presence of iRFP720 trigger mRNA. They found that constitutive expression of MCP-ADAR causes an increase in sensor activation in the absence of the trigger.

References

Gayet, R. V., Ilia, K., Razavi, S., Tippens, N. D., Lalwani, M. A., Zhang, K., ... & Collins, J. J. (2023). Autocatalytic base editing for RNA-responsive translational control. Nature Communications, 14(1), 1339. 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
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


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Parameters
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