Bio-circuitry Flashcards
Linking environment-responsive promoters to engineered gene circuits is what type of bio-circuitry?
Transcriptional.
What domains are utilised in translational bio-circuitry?
RNA aptamer domains.
What type of bio-circuitry uses protein receptors to trigger signal cascades?
Post-translational.
Operons are a key element of which type of bio-circuit?
Transcriptional.
Describe broadly the mechanism of the arsenic operon.
The arsenic repressor disassociates from the operon if it is bound by arsenite, activating the detoxifying pathway.
Why might we create a constantly-active ‘decoupled’ arsenic repressor operon?
If it is all part of the same operon, there will be constant ‘leaky’ basal activity and thus constant reporter expression.
A two-input biosensor that produces an inducer for a second biosensor is called a…?
Three-input biosensor.
Name three advantages of cell-free biosensors.
- Low cost.
- Non-GMO
- Highly stable.
- Faster response than a whole-cell biosensor.
A gate that is on with no input is?
A ‘not’ gate.
A gate that is off if either/both inputs are present is?
A ‘nor’ gate.
When combining gates, repressors can’t bind other promoters. This idea is called:
Orthogonality.
An RNA molecule with a complex secondary structure that can regulate translation is known as:
A riboswitch.
In which region of RNA are riboswitches found?
5’ untranslated region (UTR).
Binding to the _____ causes conformational changes to the riboswitch.
Aptamer region.
Name two advantages of a riboswitch as a biosensor.
- Transcription step is skipped.
- Sensor + regulator + output, all in one.
- High specificity.
What is a trans-activating riboswitch?
A riboswitch activated by another RNA molecule.
What is the example application of a trans-activating ‘toehold switch’?
Viral RNA (e.g. Ebola) could trigger the switch.
Creating an aptamer region for DNT involved trial-and-error with the addition of a bunch of random bases. True or false?
True.
Why would we want to detect DNT? Give two reasons.
Industrial pollutant + precursor to TNT
What two categories of variant can we dismiss when screening for DNT-inducable riboswitches?
- Variants that fluoresce WITHOUT any DNT (the riboswitch hasn’t folded properly)
- Variants that don’t fluoresce WITH DNT (aptamer isn’t optimised for DNT binding)
Does DNT riboswitch produce GFP? Explain.
It doesn’t produce GFP - the switch activates a protease which unlocks GFP from another protein and allows it to fluoresce.
Why create a DNT riboswitch that produces the protease to unlock GFP rather than just produce GFP?
Producing each GFP in response to DNT would be time-consuming - instead, having the GFP constantly present but dormant allows the DNT-induced protease to rapidly activate fluorescence.
What is the central advantage of protein switches over genetic circuits?
No transcription/translation events = instantaneous responses.
What are the three main types of post-translational events?
- Conformational changes.
- Protein interactions.
- Post-translation modifications (PTMs).
Name three possible choices of sensor for post-translational bio-circuits.
- Ligand binding domains.
- Metal binding motifs.
- Protein binding domains.
- Cleavage sites.
How does adding calmodulin to GDH work as a biosensor?
Calmodulin will bind to any present calcium, inducing a conformational change in the entire chimeric protein and causing a measurable increase in electron activity.
