Part:BBa_K3743011

onToehold switch

Part Description

Toehold switches are synthetic RNAs that mimic messenger RNAs whose job it is to shuttle information from the DNA to the protein-synthesizing machinery. They contain a recognition sequence (toehold) for a specific stimulus in form of a specific “input” RNA, and a recognition sequence that the protein-synthesizing machinery (ribosome) needs to bind to initiate the translation of a fused protein-coding sequence into its encoded protein product. In the absence of the “input” RNA, the toehold switch is kept in its OFF state by forming a hairpin structure that uses part of the “input” recognition sequence and the ribosome recognition sequence, which is kept inaccessible.

Usage

It can detect and bind to miRNAs from the cancerous environment whether those indicate tumor suppression or tumor growth which will give the opportunity to make decisions as in case of increased miRNAs that indicate tumor growth. A complex consists of the binding of a ribosome to the ribosomal binding site (RBS) on the Toehold switch which will destabilize the transcripts of L7Ae protein, therefore removing its inhibitory effect which allows continuous transcription at RNA levels to increase the yield of the vaccine.

Characterization by Mathematical Modelling

Blue plot achieves the maximum concentration at 1.6 after 700 seconds which represents the presence of 2 inputs in the AND (absence of TMP and binding of miRNA to toeholdOn switch upstream to the vaccine) and the red plot represents the inhibition of the transcription in absence of one of the two inputs wither the presence of the TMP so stabilized the destabilizing domain and inhibiting the circuit or absence of binding of miRNA to ToeholdOn switch

Figure 1. AND gate model simulation between TMP administration and ToeholdOn switch.

Structural Characterization

In order to model the structural stability of ToeholdOnmotif switch, we used RNAfold tool that predicts secondary structures of single stranded RNA.

The free energy of the thermodynamic ensemble is -24.52 kcal/mol.

The frequency of the MFE structure in the ensemble is 59.20 %.

The ensemble diversity is 1.76 .

Figure 2.Structural Characterization of Toehold.

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]