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<br><br>It is also known that the proteins are prone to degradation, which implies that the protein production can be ‘shielded’ by this degradation. Therefore, knowing how the degradation operates, turns out to be essential to the description of E. coding. It is also important to take into consideration that this degradation is proportional to the protein quantity; so the protein count, in controlled optimal conditions, will tend to an equilibrium at every IPTG concentration. | <br><br>It is also known that the proteins are prone to degradation, which implies that the protein production can be ‘shielded’ by this degradation. Therefore, knowing how the degradation operates, turns out to be essential to the description of E. coding. It is also important to take into consideration that this degradation is proportional to the protein quantity; so the protein count, in controlled optimal conditions, will tend to an equilibrium at every IPTG concentration. | ||
<div class="body-subtitle">Complex formation</div> | <div class="body-subtitle">Complex formation</div> | ||
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<b>Cas complex 'X'</b> | <b>Cas complex 'X'</b> | ||
− | <b>msr-msd with RT complex 'MP'</b> | + | <br><br>The key complex in the whole system is the Cas complex, responsible for the eventual message insertion in the bacteria's genome. Aiming for a simple and compact notation, this complex will hereby be denoted by ‘X’. |
+ | <br><br>The formation of X is identified in previous research as the formation of two dimers of Cas1, and one dimer of Cas2, forming a hexamer of Cas1 and Cas2. X formation is taken as the binding of 4 Cas1 proteins with 2 Cas2 proteins | ||
+ | <br><br>Although it may happen that X is stable, a backward biochemical reaction is considered as possible in order to improve the precision of the model. | ||
+ | |||
+ | <br><br><b>msr-msd with RT complex 'MP'</b> | ||
+ | |||
+ | <br><br> | ||
+ | |||
+ | <br><br> | ||
+ | <br><br> | ||
+ | <br><br> | ||
<div class="body-subtitle">Complex formation</div> | <div class="body-subtitle">Complex formation</div> |
Revision as of 16:17, 17 October 2018
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Mathematical Model
E. coding description
Introduction
The description of our system had a progressive development parallel to the realization of the whole project. Initially, a simple system of linear coupled differential equations was proposed to describe the whole system, but as the understanding of all the aspects the system involves improved, the model was continuously refined to encompass a more suitable description that became both accurate and practical.
Biological System
Biological systems modeling is the creation of a kinetics diagram that connects all the important species of the system through relationships that represent bio-chemical reactions.
In the case of the E. coding system, the methodology consisted in:
- The identification of each individual component
- Consulting literature on the possible interactions
- Creating a kinetic diagram
- Establishing the whole differential equations system via biochemical reactions and the eventual message insertion inside our bacteria
Identification
In the bateria, five precursor species were identified as the ones relevant for the system. These precursors are precisely Cas1 protein, Cas2 protein, the msr-msd (UM from ‘unprocessed message’), the retrotranscriptase (RT) protein, and IPTG (I from 'inductor'). The first four species are produced by IPTG regulated transcription, which has some advantages, mainly the fact that the bacteria is not able to metabolize it, so its concentration does not change.
Protein expression
Production
Research on IPTG induction in our plasmid led us to the Hill equation, an expression that is simple, practical and commonly used in regulated protein production modeling.
It is well known that the induction stimulates the transcription of RNA which then is translated by the ribosomes into proteins. So, the Hill equation is valid when it is assumed that, as soon as the RNA is produced, it is immediately processed and translated into the corresponding proteins by the bacteria.This means that all these four species are able to be modeled by the Hill equation.
An important thing to consider is the native production of these proteins by the bacteria. Although it is expected that this basal production is insignificant when compared to the IPTG regulated production, the use of this basal production increases the accuracy in which the insertions are related to the inductor concentrations.
Degradation
It is also known that the proteins are prone to degradation, which implies that the protein production can be ‘shielded’ by this degradation. Therefore, knowing how the degradation operates, turns out to be essential to the description of E. coding. It is also important to take into consideration that this degradation is proportional to the protein quantity; so the protein count, in controlled optimal conditions, will tend to an equilibrium at every IPTG concentration.
Complex formation
Cas complex 'X'
The key complex in the whole system is the Cas complex, responsible for the eventual message insertion in the bacteria's genome. Aiming for a simple and compact notation, this complex will hereby be denoted by ‘X’.
The formation of X is identified in previous research as the formation of two dimers of Cas1, and one dimer of Cas2, forming a hexamer of Cas1 and Cas2. X formation is taken as the binding of 4 Cas1 proteins with 2 Cas2 proteins
Although it may happen that X is stable, a backward biochemical reaction is considered as possible in order to improve the precision of the model.
msr-msd with RT complex 'MP'
Complex formation
Complex interactions
Insertion
Differential Equations System
Kinetic diagram
Equations
Parameters
Modified Quasi-Steady State Approximation
Equations
Parameters
Protein Production characterization
Protein Production Model
Equations
Parameters
Predictions
Sudden bulk induction
Expression
Insertions
Initial conditions expansion
Industry induction
Expression
Insertions
Initial conditions expansion