Difference between revisions of "Team:BIT-China/Model/GPCRpathway"

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    <section class="content_container" id="mytop">
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    <h2 class="title-h2">MODEL-Docking </h2>
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    <div class="cd-section" id="Purpose">
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<h3 class="title-h3">STRUCTURE MODEL</h3>
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<h4 class="title-h4">Purpose </h4>
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<p class="my-content-p">In order to confirm whether this "radar" T1R2/T1R3 can "sense" the sweetness of different sweetener, we simulated the model of receptor's structure[1] [4]. It was helpful to observe how the sweeteners binding to T1R2/T1R3 receptor visually. Moreover, we expected to find some unknown sweeteners binding sites based on this model, even some ideal sweeteners are still unknown. </p>
  
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<h4 class="title-h4">Methods</h4>
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<p class="my-content-p">To make the signal input more accurate and reliable, we simulated the T1R2/T1R3 receptor's structure model through SWISS-MODEL. Meanwhile, according to Chemdraw 2D and Chemdraw 3D, we constructed some sweetners' model. And the docking process was performed by using Autodock Vina.</p>
  
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<section class="content_container" id="mytop">
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<h4 class="title-h4">Results</h4>
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<div class="section-upline cd-section" id="Purpose">
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<p class="my-content-p">We used homology modeling to obtain the structure of human sweet receptor T1R2/T1R3. According to the crystal protein structure of similar receptor in mice, we only simulated ligand-binding-domain of our receptor. </p>
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  <h2 class="title-h2">MODEL-GPCR Pathway </h2>
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<div class="my-content-box">
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<h3 class="title-h3">Purpose</h3>
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              <img class="formula50" src="https://static.igem.org/mediawiki/2017/6/65/T--BIT-China--2017modeling_pic3.png" />
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<p class="my-content-p">By establishing the signal transduction model, we can intuitively understand how the sweetener signal conducts in the GPCR and the details of each step of the signal arrival thorough, which provides a powerful help for our transformation of the pathway, amplification of the signal, while the simulation results can also give us the future design of biological sweetness meter to provide theoretical help.</p>
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              <span>Fig. 3 The simulated structure of human sweetness receptors' ligand-binding domain (LBD)</span>
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</div>
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            </div>
  
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<div class="section-upline cd-section" id="Method">
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<p class="my-content-p">Then, taking advantage of software Chemdraw 2D and Chemdraw 3D, we established some natural or artificial sweeteners' structure.</p>
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<h3 class="title-h3">Method</h3>
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<p class="my-content-p">Our GPCR model based on three presuppositions:
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<ul>
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<li class="my-content-li">1. We assume that T1R2-T1R3 receptor does not have synergistic effect.</li>
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<li class="my-content-li">2. We hypothesize that the combination between s1</li>
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<li class="my-content-li">3. We assume that the combination of the rate constant and the initial concentration of the sweeteners and alpha pheromone are consistent.</li>
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</ul>
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</p>
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<p class="my-content-p">We establish the equations among variable quantities based on Mass-action law; and we use ordinary differential equations (ODEs) that can be solved by MATLAB. We remake the pheromone signal system in yeast into four modules: T1R2-T1R3 receptor activation, G-protein cycle activation, the MAPK cascade, and gene expression.
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<p class="my-content-p">
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<p class="my-content-p">T1R2-T1R3 receptor activation part:</p>
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<img class="formula50" src="https://static.igem.org/mediawiki/2017/5/50/T--BIT-China--2017modeling_pic4.png" />
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<img class="formula2"src="https://static.igem.org/mediawiki/2017/3/3f/BIT-China_model-GPCR_fig1.png"/>
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<span>Fig. 4 Three dimensional structure of some sweeteners</span>
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            </div>
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<p class="my-content-p">Docking process was carried out under Autodock Vina (Fig. 5-7).</p>
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<div class="my-content-box">
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<p class="my-content-p">reaction diagram</p>
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<p class="my-content-p">It can be modeled in a set of differential equations: </p>
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<img class="formula" src="https://static.igem.org/mediawiki/2017/f/fa/BIT-China_model-GPCR_formula1.png"/>
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</div>
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<div class="my-content-box">
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<p class="my-content-p">Rate equations:</p>
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<img class="formula" src="https://static.igem.org/mediawiki/2017/7/76/BIT-China_model-GPCR_formula2.png"/>
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</div>
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<div class="my-content-box">
 
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<p class="my-content-p">G-protein cycle activation part:</p>
 
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<img class="formula2" src="https://static.igem.org/mediawiki/2017/4/45/BIT-China_model-GPCR_fig2.png"/>
 
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</div>
 
  
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<div class="my-content-box">
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<div class="my-img-box" style="align-items: flex-end; justify-content: space-around;">
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<p class="my-content-p">It can be modeled in a set of differential equations:</p>
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<img style="width: 30%; height: 30%" src="https://static.igem.org/mediawiki/2017/7/7b/T--BIT-China--2017modeling_pic5.png" alt="">
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<img class="formula" src="https://static.igem.org/mediawiki/2017/7/74/BIT-China_model-GPCR_formula3.png"/>
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<img style="width: 30%; height: 30%" src="https://static.igem.org/mediawiki/2017/5/54/T--BIT-China--2017modeling_pic6.png" alt="">
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<img style="width: 25%; height: 25%" src="https://static.igem.org/mediawiki/2017/7/7c/T--BIT-China--2017modeling_pic7.png" alt="">
 
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<div class="my-img-box">
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<span>Fig. 5． Docking result of different sweeteners </span>
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<p class="my-content-p">Rate equations:</p>
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<span>Fig. 6． Docking result of aspartame </span>
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<img class="formula" src="https://static.igem.org/mediawiki/2017/1/14/BIT-China_model-GPCR_formula4.png"/>
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<span>Fig. 7． Docking result of stevioside </span>
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<div class="my-content-box">
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<p class="my-content-p">The MAPK cascade part:</p>
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<img class="formula2" src="https://static.igem.org/mediawiki/2017/7/70/BIT-China_model-GPCR_fig3.png"/>
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</div>
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<div class="my-content-box">
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<p class="my-content-p">It can be modeled in a set of differential equations:</p>
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<img class="formula" src="https://static.igem.org/mediawiki/2017/0/04/BIT-China_model-GPCR_formula5.png"/>
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<div class="my-content-box">
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<p class="my-content-p">Rate equations:</p>
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<img class="formula" src="https://static.igem.org/mediawiki/2017/4/4c/BIT-China_model-GPCR_formula6.png"/>
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<div class="my-content-box">
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<p class="my-content-p">Gene expression part:</p>
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<img class="formula2" src="https://static.igem.org/mediawiki/2017/9/98/BIT-China_model-GPCR_fig4.png"/>
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</div>
 
</div>
  
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<div class="my-content-box">
 
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<p class="my-content-p">It can be modeled in a set of differential equations:</p>
 
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<img class="formula" src="https://static.igem.org/mediawiki/2017/d/d8/BIT-China_model-GPCR_formula7.png"/>
 
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</div>
 
  
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<div class="my-content-box">
 
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<p class="my-content-p">Rate equations</p>
 
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<img class="formula" src="https://static.igem.org/mediawiki/2017/1/14/BIT-China_model-GPCR_formula8.png"/>
 
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<table class="table-co">
 
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            <caption>This table gives the various parameter values we found in literature:</caption>
 
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<div class="section-upline cd-section" id="PSR">
 
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<h3 class="title-h3">Primary simulation result</h3>
 
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<div class="my-content-box">
 
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<img class="formula2" src="https://static.igem.org/mediawiki/2017/2/20/BIT-China_model-GPCR_fig5.png"/>
 
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<p class="my-content-p">The result is calculated by our initial model. As we can see, RFP fluorescence intensity will saturate when sweetener’s concentration reaches 2uM. This result is consistent with what we see from the literature.</p>
 
  
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<p class="my-content-p">So we transform our system based on this model, the results are as follows;</p>
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<h4 class="title-h4">Discussion</h4>
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<div class="my-content-box">
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<p class="my-content-p">According to the results, all common, representative sweet substances can bind to human T1R2/T1R3 receptors on ligand-binding Domain (LBD). That means there are a lot of sites in receptor can provide advantage of suitable binding. Although model is only a part of the whole receptor, this domain is the most important domain for binding. It proves that this receptor can "taste" the sweetness and our project has enormous potential to be used in a wide range of sweet material testing compared to E-Tong.</p>
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<img class="formula2" src="https://static.igem.org/mediawiki/2017/6/64/BIT-China_model-GPCR_fig6.png"/>
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<p class="my-content-p">Our docking results estimated that ligand-binding domain (LBD) of human T1R2/T1R3 receptors could bind with almost all kinds of known sweeteners. It provided evidence that this receptor has an ability to "taste" the sweetness. Meanwhile, our results also helped us to speculate that the number of both binding-site and binding-molecule also might influence the sense of sweetness.</p>
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</div>
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<p class="my-content-p">However, our model still needs optimization. We plan to enhance the docking model and the more professional direct would be supplied by it.</p>
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<p class="my-content-p">Based on the experimental data and the result of simulation we can see that the concentration of 5-10 uM works best. Our analysis concludes that we have not yet achieved the results of yeast transformation.</p>
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</div>
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<p class="my-content-p">At the same time, we also find the situation of different sweeteners combination is different, for example the binding number, binding concentration and so on, which demonstrates people for the perception of sweet substances not only depends on the sweetness of the sweetener, but also on the concentration and the number of binding. In the future, it can provide the guidance for us to develop sweetness standards.</p>
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<div class="section-upline cd-section" id="Discussion">
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<h3 class="title-h3">Discussion</h3>
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<p class="my-content-p">It is clear that the results of our initial model and experimental model are different, indicating that our model is not accurate enough. We need to keep working, such as changing the value of certain parameters, rewriting some important equations, so that our model and our experimental can combine perfectly.</p>
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<p class="my-content-p">But our model needs to be improved, in the future, we will use molecular dynamics modeling (molecular docking) and other methods to modify the sweet receptor model utilizing software Gauss View through the Gaussian force field and other mechanical system. The appropriate deformation of the small molecular material is also going to be simulated for making docking results be closer to the real situation.</p>
 
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<div class="section-upline cd-section" id="References">
 
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<h3 class="title-h3">References</h3>
 
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<p class="my-content-p">1.Grant E. DuBois, Molecular mechanism of sweetness sensation, Physiology & Behavior 164 (2016) 453–463.</p>
 
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<p class="my-content-p">2. Bente Kofahl1 and Edda Klipp2, Modelling the dynamics of the yeast pheromone<
 
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pathway, Published online in Wiley InterScience, Yeast 2004; 21: 831–850.</p>
 
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Revision as of 14:10, 27 October 2017

BIT-CHINA

MODEL-Docking

STRUCTURE MODEL

Purpose

In order to confirm whether this "radar" T1R2/T1R3 can "sense" the sweetness of different sweetener, we simulated the model of receptor's structure[1] [4]. It was helpful to observe how the sweeteners binding to T1R2/T1R3 receptor visually. Moreover, we expected to find some unknown sweeteners binding sites based on this model, even some ideal sweeteners are still unknown.

Methods

To make the signal input more accurate and reliable, we simulated the T1R2/T1R3 receptor's structure model through SWISS-MODEL. Meanwhile, according to Chemdraw 2D and Chemdraw 3D, we constructed some sweetners' model. And the docking process was performed by using Autodock Vina.

Results

We used homology modeling to obtain the structure of human sweet receptor T1R2/T1R3. According to the crystal protein structure of similar receptor in mice, we only simulated ligand-binding-domain of our receptor.

Fig. 3 The simulated structure of human sweetness receptors' ligand-binding domain (LBD)

Then, taking advantage of software Chemdraw 2D and Chemdraw 3D, we established some natural or artificial sweeteners' structure.

Fig. 4 Three dimensional structure of some sweeteners

Docking process was carried out under Autodock Vina (Fig. 5-7).

Fig. 5． Docking result of different sweeteners Fig. 6． Docking result of aspartame Fig. 7． Docking result of stevioside

Discussion

According to the results, all common, representative sweet substances can bind to human T1R2/T1R3 receptors on ligand-binding Domain (LBD). That means there are a lot of sites in receptor can provide advantage of suitable binding. Although model is only a part of the whole receptor, this domain is the most important domain for binding. It proves that this receptor can "taste" the sweetness and our project has enormous potential to be used in a wide range of sweet material testing compared to E-Tong.

Our docking results estimated that ligand-binding domain (LBD) of human T1R2/T1R3 receptors could bind with almost all kinds of known sweeteners. It provided evidence that this receptor has an ability to "taste" the sweetness. Meanwhile, our results also helped us to speculate that the number of both binding-site and binding-molecule also might influence the sense of sweetness.

However, our model still needs optimization. We plan to enhance the docking model and the more professional direct would be supplied by it.

At the same time, we also find the situation of different sweeteners combination is different, for example the binding number, binding concentration and so on, which demonstrates people for the perception of sweet substances not only depends on the sweetness of the sweetener, but also on the concentration and the number of binding. In the future, it can provide the guidance for us to develop sweetness standards.

But our model needs to be improved, in the future, we will use molecular dynamics modeling (molecular docking) and other methods to modify the sweet receptor model utilizing software Gauss View through the Gaussian force field and other mechanical system. The appropriate deformation of the small molecular material is also going to be simulated for making docking results be closer to the real situation.

TOP

Contact

Institute of Biotransformation and synthetic biosystem

School of life science

Beijing Institute of Technology

100081, Beijing

Email: lichun@bit.edu.cn