Team:Bielefeld-Germany/Modell
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- | The next step will be, to add more substrates an different laccases to our model. Therefore we will need mot data about laccases. K<sub>cat</sub> K<sub>M</sub><sup>-1</sup> values has to be determined and wich laccase preferes wich substrate. Additionaly we have the opportunity to work with a lab sewageplant. So we´re going to test our model in defined conditions. | + | The next step will be, to add more substrates an different laccases to our model. Therefore we will need mot data about laccases. K<sub>cat</sub> K<sub>M</sub><sup>-1</sup> values has to be determined and wich laccase preferes wich substrate. Additionaly we have the opportunity to work with a [https://2012.igem.org/Team:Bielefeld-Germany/Sewage_treatment_plant#Conclusion lab sewageplant]. So we´re going to test our model in defined conditions. |
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Revision as of 00:21, 27 September 2012
Contents |
Model of a fixed-bed reactor
In our project we plan to construct a fixed-bed reactor, where immobilized laccases degrade synthetic estrogen and other harmful substances. As a small selection we plan to characterize our different laccases for three estrogens, three analgesics, four PAH´s, one insecticide and three possible redox mediators. If we could model the degradation of one substrate by one laccase, we could easily replace the specific Kcat KM-1 quotient of other laccases and the amounts of the other substrates.
We ignore the possible cofactors ABTS, syringaldazine and viuloric acid, because on the one hand using these cofactors would increase the costs, and on the other hand they are harmful substances. As shown by Team Substrate Analytic TVEL0 degrades ethinyl estradiol without a redox mediator. So the reaction should follow the michaelis menten kinetics.
The reaction
The normal michaelis menten cinetic (1) can be adaptet for low substrate concentrations. In order to do this formula (2) replaces [ES] in formula (3). The result is formula (4). The velocity of this reaction describes the cange of the substrate concentration over time (5). Formula (6) and (7) steps to solve the integral. (8) is the resulting formula to determine the time dependent substrate concentration by a given startconcentration A.
This transformation can be found in "Stryer biochemie". This formula is suitable for very low substrate concentrations. In this case we can estimate the substrate concentrations below 0.1 µg L-1.
Because we can not find any information about the Kcat KM-1 quotient for the degradation of estradiol in the enzyme database [http://www.brenda-enzymes.info Brenda] we decided to use different Kcat KM-1 quotients for the degradation of [http://www.sigmaaldrich.com/catalog/product/sigma/a1888?lang=de®ion=DE ABTS], a red ox mediator of the laccase. In the following picture we show a few possible reactions of our laccase.
We use Kcat KM-1 values fro 11 to 643500 and get degradation time points from 0,07s to 1500s. Furthermore we try to integrate different temperatures to our model. In a span from 10°C to 30°C and a Kcat KM-1 value of 1, the degradation differs from 40s to 100s.
Extend the model with sewageplant data
We got discharge water data from Schloss Holte about the water tempreture, pH value and flowrate. The temperature and pH value have a direct influence of enzymatic activity. Dependent on the enzymatic activity we want to calculate the time our fixed-bed reactor will need to degrdate 80 % of the substrates. This will be the dewll time. Combined with the actual flowrate we can determine the reactorsize. To estimate the feasibility we wart to know how much enzyme has to be produced for a wewage plant. A 3D model of the requirde enzme amount dependent of Kcat KM-1 values and flowrate.
Outlook
The next step will be, to add more substrates an different laccases to our model. Therefore we will need mot data about laccases. Kcat KM-1 values has to be determined and wich laccase preferes wich substrate. Additionaly we have the opportunity to work with a lab sewageplant. So we´re going to test our model in defined conditions.
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