IGEM:IMPERIAL/2006/project/Oscillator/project browser/Test Killing Predator Construct

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Super Parts Predator Construct
Actual Part AiiA Testing Construct <bbpart>J37022</bbpart>
Sub Parts <bbpart>R0010</bbpart> <bbpart>B0034</bbpart> <bbpart>J37023</bbpart> <bbpart>B0015</bbpart>


Use content from: IGEM:IMPERIAL/2006/project/parts/BBa_J37022

Motivation

AiiA forms an integral part of the biological oscillator acting as our predator in the Lotka-Volterra Predator-Prey Model. By testing and characterising this part individually, we can find the rate at which AiiA converts N-acyl homoserine lactone (AHL) into it's inactive form by cleaving the lactone ring (AHL-lactonase activity) at the ester bond.

Parameters to explore:

  • Activity of AiiA
  • Half-life of AiiA

Michaelis-menten kinetics would determine the activity of the enzyme, thus if we can characterise AiiA based upon it's activity (turnover rate) and half-life, the AiiA construct (part J37025) can be placed anywhere and easily integrated into more complex systems given it's transfer function defined by the enzyme parameters.

Part Interface

Testing of this construct involves two stages.

Stage 1: To produce AiiA by inducing transcription by adding IPTG.

Stage 2: The second stage is to degrade AHL with AiiA to measure the actual enzyme activity.

By characterising stage one (getting a transfer function for a input of IPTG and output of AiiA), we will be able to decouple the two stages so we will not need to measure the AiiA concentration in between the two stages. We can get a calibration curve knowing that for a given amount of IPTG we use to induce transcription, we will be obtaining some concentration of AiiA, within a given range.

INPUTS OUTPUTS
IPTG AiiA

Our second stage can be summarised as follows:

INPUTS OUTPUTS
AiiA [math]\displaystyle{ \frac{d[AHL]}{dt} }[/math]

Expected behavior and performances

Stage 1 Intefrace Specifications (IPTG to AiiA)

Characteristics IPTG AiiA
Range [0 mM, 10 mM] Unknown
Tolerance Unknown Unknown
Transfer function n/a [math]\displaystyle{ [AiiA] = \frac{V_{max}}{\delta_{deg}} \times \frac{[IPTG]}{K_m + [IPTG]} }[/math]
Latency

Stage 2 Intefrace Specifications (AiiA to d[AHL])

Characteristics AiiA d[AHL]
Range Unknown Unknown
Tolerance Unknown Unknown
Transfer function n/a [math]\displaystyle{ v = \frac{k_2[AiiA][AHL]}{K_m + [AHL]} }[/math]
Latency
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