Team:University College London/Module 1/Modelling

From 2012.igem.org

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| R5 || POPexmRNANahr.Psal → GFP.mRNA|| 0.11 || Transcription rate of GFP in molecules/sec (for GFP size 720bp<sup>10</sup>, transcription rate in E.coli 80bp/sec<sup>8</sup>)
| R5 || POPexmRNANahr.Psal → GFP.mRNA|| 0.11 || Transcription rate of GFP in molecules/sec (for GFP size 720bp<sup>10</sup>, transcription rate in E.coli 80bp/sec<sup>8</sup>)
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| R6 || GFP.mRNA → GFP|| 0.04 || Translation rate of GFP in molecules/sec (for laccase size 240aa<sup>10</sup>, translation rate in E.coli 20aa/sec<sup>8</sup>)
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| R6 || GFP.mRNA → GFP|| 0.04 || Translation rate of GFP in molecules/sec (for GFP size 240aa<sup>10</sup>, translation rate in E.coli 20aa/sec<sup>8</sup>)
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| R11 || Lin → Lindegp || 0.03 || Degradation rate of laccase<sup>11</sup> must be taken into account due to suboptimal conditions
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| R9 || GFP.mRNA → 0 || 0.03 || Degradation rate of GFP mRNA product<sup>11</sup> must be taken into account due to suboptimal conditions
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| R7 || Lin → Lex || Forward: 0.9 <br /> Backward: 0.1 || Our laccase is a periplasmic enzyme, therefore most of it is released in the periplasm.  However, we assumed leakage of 20 percent based on suboptimal conditions in the periplasm<sup>12</sup>, which is able to degrade polyethene.
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== Results ==
== Results ==
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We ran three simulations in SimBiology, each over a different timespan:
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[[File:detectiongraph.jpg]]
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[[File:deg1.png]] In the first second of laccase production, we see polyethylene degradation beginning from around 0.6 sec (Outside2.PEdegp)
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[[File:deg2.png]] After 10 seconds, the first few molecules have been degraded. 
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[[File:deg3.png]] At 100 seconds, the rate of degradation has risen to almost 1 PE molecule per second.
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From these results, we can conclude that
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== References ==
== References ==
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9. Park H, Lim W, Shin H (2005) In vitro binding of purified NahR regulatory protein with promoter Psal. <i>Biochimica et Biophysica Acta</i> 1775: 247-255
9. Park H, Lim W, Shin H (2005) In vitro binding of purified NahR regulatory protein with promoter Psal. <i>Biochimica et Biophysica Acta</i> 1775: 247-255
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10. Laccase size: http://partsregistry.org/Part:BBa_K729002
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10. http://partsregistry.org/wiki/index.php?title=Part:BBa_I13522
11. To follow
11. To follow
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12. Young K, Silver LL (1991) Leakage of periplasmic enzymes from envA1 strains of Escherichia coli. <i>J Bacteriol.</i> 173: 3609–3614
 
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13. To follow (c208 ref??)
 
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14. 5. Ding Z, Peng L, Chen Y, Zhang L, Gu Z, Shi G, Zhang K (2012) Production and characterization of thermostable laccase from the mushroom, Ganoderma lucidum, using submerged fermentation.  <i>African Journal of Microbiology Research</i> 6: 1147-1157.  DOI: 10.5897/AJMR11.1257
 
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[[File:detectiongraph.jpg]]
 
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1. GFP size: http://partsregistry.org/wiki/index.php?title=Part:BBa_I13522
 
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2. NahR size: http://www.xbase.ac.uk/genome/azoarcus-sp-bh72/NC_008702/azo2419;nahR1/viewer
 
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Revision as of 15:04, 21 September 2012

Contents

Module 1: Detection

Description | Design | Construction | Characterisation | Modelling | Conclusions

Modelling

As our detection module is so closely tied up with our degradation module, our cell model for this module examines the hypothetical expression of GFP on detection of POPs. This allows us to assess the efficacy of detection as a stand-alone module before we look at its performance when incorporated into our system.

Detectionnet.jpg

Note that this model is very similar to our cell model for degradation. This model, however, looks only at detection response - modelled here by GFP production - and ignores the production and action of laccase.

-External represents the constant association and disassociation of persistent organic pollutants (POPs) that takes place in the ocean.

-Cell represents the reactions taking place inside the cell

Species

Species Initial value (molecules) Notes
PE 0.044 Polyethylene found in North Pacific Gyre (value per cubic metre)1,2
POPex 0.0 Persistent organic pollutants (ex = extracellular) that are not adhered to plastic surface
PEPOPex 9.24E-5 Persistent organic pollutants (ex = extracellular) that are adhered to the plastic surface3
POPin 0.5 Persistent organic pollutants (in = intracellular) assumed from E. coli membrane permeability 4
mRNANahR 0.0 NahR mRNA product
POPinNahR 0.0 Complex of the above two molecules
POPinNahRpSal 0.0 Complex of the above molecule and pSal (promoter that induces laccase transcription)
GFP 0.0 Green fluorescent protein
GFPmRNA 0.0 GFP mRNA product

Reactions taking place in the model

Number Reaction Reaction rate (molecules/sec) Notes
R1 PE + POPex ↔ PEPOPex Forward: 1000
Backward: 1 		Pops have 1000 to 10000 times greater tendency to adhere to plastic than float free in the ocean5
R2 POPex ↔ POPin Forward: 0.6
Backward: 0.4 		Based on membrane permeability4: diffusion gradient
R3 POPin + mRNA.Nahr → POPin.mRNA.Nahr Forward: 1
Backward: 0.0001 		Based on the assumption that the chemical structure/size of POPs is similar to salycilate6. Salycilate binds to the NahR mRNA product, which complex then binds to the pSal promoter.
R7 0 → mRNA.Nahr Forward: 0.088
Backward: 0.6 		Transcription rate of NahR in molecules/sec (for NahR size 909 bp7, transcription rate in E.coli 80bp/sec8) under constitutive promoter control
R4 POPinmRNANahr → POPinmRNANahr.Psal Forward: 78200
Backward: 0.191 9 		NahR to pSal binding based on the assumption that POP-NahR binding has no effect on NahR-pSal binding
R5 POPexmRNANahr.Psal → GFP.mRNA 0.11 Transcription rate of GFP in molecules/sec (for GFP size 720bp10, transcription rate in E.coli 80bp/sec8)
R6 GFP.mRNA → GFP 0.04 Translation rate of GFP in molecules/sec (for GFP size 240aa10, translation rate in E.coli 20aa/sec8)
R9 GFP.mRNA → 0 0.03 Degradation rate of GFP mRNA product11 must be taken into account due to suboptimal conditions

Results

Detectiongraph.jpg

References

1. Goldstein M, Rosenberg M, Cheng L (2012) Increased oceanic microplastic debris enhances oviposition in an endemic pelagic insect, Biology Letters 10.1098

2. Andrady AL (2011) Microplastics in the marine environment. Marine Pollution Bulletin 62: 1596-1605

3. To follow

4. To follow

5. Mato Y, Isobe T, Takada H, Kanehiro H, Ohtake C, Kaminuma T (2001) Plastic Resin Pellets as a Transport Medium for Toxic Chemicals in the Marine Environment. Environ. Sci. Technol. 35: 318-324

6. https://2011.igem.org/Team:Peking_S/project/wire/harvest

7. http://www.xbase.ac.uk/genome/azoarcus-sp-bh72/NC_008702/azo2419;nahR1/viewer

8. http://kirschner.med.harvard.edu/files/bionumbers/fundamentalBioNumbersHandout.pdf

9. Park H, Lim W, Shin H (2005) In vitro binding of purified NahR regulatory protein with promoter Psal. Biochimica et Biophysica Acta 1775: 247-255

10. http://partsregistry.org/wiki/index.php?title=Part:BBa_I13522

11. To follow


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