Team:PennState

From 2008.igem.org

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  <img src="https://static.igem.org/mediawiki/2008/d/df/Penn_state_igem_logo.JPG" alt="Penn State" />
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     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState" >Home</a> </td>
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     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState" title="Welcome!">Home</a> </td>
     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState/Team" >The Team</a> </td>
     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState/Team" >The Team</a> </td>
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     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState/Project" >The Project</a> </td>
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     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState/Project" title="Full Abstracts.">The Project</a> </td>
     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState/Parts">Parts</a> </td>
     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState/Parts">Parts</a> </td>
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     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState/Modeling" >Modeling</a> </td>
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     <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState/Notebook" title="Day to day lab activity">Notebook</a> </td>
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    <td align="center" ><a class="mainLinks" href="https://2008.igem.org/Team:PennState/Notebook" >Notebook</a> </td>
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  <h4 style="margin-top: 0;">Hormone Biosensors</h4>
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  <h4>Diauxie Elimination</h4>
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   <dd><a href="hbintro" title="Intro to Endocrine Disruption, pseudoestrogens, pthalates, nuclear hormone receptors, and our goals">Introduction</a></dd>
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   <dd><a href="https://2008.igem.org/Team:PennState/diauxie/intro">Introduction</a></dd>
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  <dt>Smart Fold</dt>
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   <dd><a href="https://2008.igem.org/Team:PennState/diauxie/TheSystem">The System</a></dd>
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   <dd><a href="https://2008.igem.org/Team:PennState/diauxie/Strategies">Strategies</a></dd>
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   <dd><a href="smartfold/parts" title="Parts submitted to the registry for this project">Parts</a></dd>
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   <dd><a href="https://2008.igem.org/Team:PennState/diauxie/progress">Progress</a></dd>
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   <dd><a href="smartfold/references">References</a></dd>
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   <dd><a href="https://2008.igem.org/Team:PennState/diauxie/conclusions">Conclusions</a></dd>
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  <dt>Nuclear Fusion</dt>
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   <dd><a href="https://2008.igem.org/Team:PennState/diauxie/references">References</a></dd>
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  <h4><acronym title="Nuclear Hormone Receptor">NHR Biosensors</acronym><br/></h4>
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   <dd><a href="diauxie/intro">Introduction</a></dd>
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   <dd><a href="https://2008.igem.org/Team:PennState/NHR/introduction">NHR Introduction</a></dd>
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   <dd><a href="diauxie/overview">Overview</a></dd>
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   <dd><a href="diauxie/parts" title="Parts submitted to the registry for this project">Parts</a></dd>
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   <dd><a href="https://2008.igem.org/Team:PennState/smartfold/overview">Phthalate Biosensor</a></dd>
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  <dd><a href="diauxie/references">References</dd>
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   <dd><a href="https://2008.igem.org/Team:PennState/fusion/overview">BPA Biosensor</a></dd>
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<td valign="top" id="pagecontent"><span style="font-size: 16pt">PENN STATE iGEM 2008</span>
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<td valign="top" id="pagecontent" width="80%"><span style="font-size: 16pt">Welcome to Penn State iGEM 2008</span>
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<p>Welcome to the Penn State iGEM 2008 team’s website.  We are currently working hard at a few different projects for this year's competition. In early May we began brainstorming and came up with a couple of ideas to create biosensors that use human nuclear hormone receptors to recognize potentially harmful ligands. These receptor systems occur naturally in the human body, but our goal is to retain and utilize their functions in <em>Escherichia Coli</em>. We are also finishing up one of last year's projects which is aimed at creating a more efficient bioproduction process by altering how <em>E. Coli</em> selects between the utilization of 5 and 6 carbon sugars.  Please explore our website to find out more about us and our projects!</p>
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  <p class="start"><a href="http://www.psu.edu" title="Pennsylvania State University" target="_blank">Penn State University</a> has participated in the International Genetically Engineered Machines (iGEM) competition for four years as of 2008, and we are excited to participate once more. Our team consists of 5 undergraduate students, one visiting undergrad and one high school student who work independently, coordinated through weekly meetings with our advisers. Check out our <a href="https://2008.igem.org/Team:PennState/Team" title="Penn State's 2008 iGEM team">Team Page</a> to meet us!</p>
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<p> If there are any questions or comments about the information on this site please contact us at
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  <p><strong>Our main focus for 2008 was the elimination of <a href="https://2008.igem.org/Team:PennState/diauxie/intro">diauxie</a> in <em>E. coli</em> to create a xylose inducible system independent of glucose regulation.</strong> This system could be used for creating more efficient bioproduction by altering the utilization of 5 and 6 carbon sugars. Please check out the links on the left to navigate through our work.</p>
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<a href="mailto:gjt5001@psu.edu" title="email us">gjt5001@psu.edu</a>. </p>
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  <p>Two other projects begun this year focus on creating <a href="https://2008.igem.org/Team:PennState/NHR/introduction"What kind of 'Biosensors' are we talking about?">biosensors</a> that use human nuclear hormone receptors to recognize potentially harmful compounds. These receptor systems occur naturally in the human body, but our goal is to retain and utilize their functions in <em>E. coli</em>. The links on the left should introduce you to our thought process and progress on each of these projects, and provide a fuller introduction to the topic.</p>
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  <p> Please explore our website to find out more about us and our projects! If there are any questions or comments about the information on this site please feel free to contact us at <a href="mailto:gjt5001@psu.edu" title="email us">gjt5001@psu.edu</a>. </p>
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        <tr><td style="padding-top:30px; padding-right:30px" valign="top" width="90%"><span style="font-size:14pt">Hormone Prescreening <em>E. coli</em></span>
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      <p>Two of our projects aim to construct biosensors which will ultimately serve as a water prescreening tool.  The focus of these biosensors will be to detect phthalate compounds utilizing the Peroxisome Proliferator Activated Receptor (PPAR) and detecting Bisphenol A (BPA) by the Estrogen Receptor (ER).  Recent studies show phthalates cause negative health effects such as damage to the liver and kidneys and birth defects in rodents.  Phthalates are introduced into our environment by their use as plastisizers for materials ranging from polyvinyl chloride to nail polish to small toys.  BPA is also found in plastics but instead it is used for the synthesis of hard plastics.  Once BPA enters the human body it is confused for estrogen and parallels the effects of estrogen after attaching to the ligand binding region of the ER.  Analytical detection methods for water contamination are compound specific and very costly.  Having a simple and cheap tool to screen for phthalates or BPA as a general class of compounds would eliminate the cost and time involved in detection.</p>
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      <p> We are using two of the natural human nuclear hormone receptor proteins that recognize a large class of ligands, and attempting to express them heterologously in <em>E. Coli</em>.  The complexity of this mammalian protein makes it difficult to express it in a prokaryote.  We have two different strategies to express and use these receptors to detect compounds in <em>E. Coli</em>.  </p>
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    <span style="font-size: 14pt"><a href="https://2008.igem.org/Team:PennState/MedalChecklist">Medal Checklist</a></span>
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          <td style="padding-top:30px; padding-right:30px" valign="top" width="45%"><span style="font-size: 14pt">Smart Fold Reporter</span>
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            <p> The human PPAR has three different types α, β, and γ but only two show any affect by phthalates.  We are using the alpha form which is expressed in the liver, kidney, heart, muscle, adipose tissue, and others.  There are different regions associated with nuclear hormone receptors: N-terminal, DNA binding domain (DBD), Hinge, Ligand binding domain (LBD), and C-terminal.  The LBD is the region that attracts and holds the ligand of interest.  After ligand binding the receptor usually will form a dimer, in our case PPAR will combine with Retinoid X Receptor (RXR) to form a heterodimer.  The RXR protein functions much like the PPAR but in this case it does not need to attach a ligand before dimerization.  The heterodimer will bind to Peroxisome Proliferator Response Element (PPRE) via the DBD and activates transcription.  Most often a coactivator complex is required for transcriptional activation which involves proteins SRC-1, CBP and others.  </p>
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    <h3>Related Links</h3>
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      <li><a href="http://partsregistry.org/Main_Page" title="Parts Registry">Parts Registry</a></li>
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<p>This <em>Smart Fold Reporter</em> project uses altered growth conditions so that the entire PPAR protein is successfully expressed and used to transcriptionally report for the presence of phthalates. Expressing the entire PPAR in E. Coli has proven difficult which could be caused by toxicicity to the cells from the DBD. To overcome this problem we are going to treat the E. Coli with carbonyl cyanide m-chlorophenyl-hydrazone (CCCP) which is an uncoupler of oxidative phosphorylation.  This strategy would correlate to the heat shock proteins involved with synthesis in the human body.  The cells that have the PPAR plasmid will be grown on plates containing Timentin which prevents growth of bacteria without plasmid. The expression of the PPAR and RXR also need tight requlation so the arabinose operon will be used.  A green fluorescent protein will be placed after the PPRE to signal transcription after heterodimer binding.
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<li><a href="http://www.che.psu.edu/Faculty/Cirino/" title="Prof. Pat Cirino's Lab website" target="_blank">Prof. Pat Cirino's Lab website</a></li>
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<li><a href="http://www.abe.psu.edu/fac/Richard/Overview.htm" title="Prof. Tom Richard's Lab website" target="_blank">Prof. Tom Richard's Lab website</a></li>
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          <td style="padding-top:30px; padding-right:30px" valign="top" width="45%"><span style="font-size:18px">Nuclear Fusion</span>
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<li><a href="http://openwetware.org/wiki/IGEM:PennState" title="Penn State iGEM OpenWetWare" target="_blank">Penn State iGEM OpenWetWare</a></li>
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<img src="https://static.igem.org/mediawiki/2008/d/d9/PSU2008iGEM_BPAimage.png" alt="[img]" style="float:left; margin:5px;width: 30%;"/>
 
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            <p> The <em>Nuclear Fusion</em> project currently has two similar directions that it may turn but both involve a plasmid construct very generously donated to our iGEM team from David W. Wood, Department of Chemical Engineering at Princeton University.  Research in their lab has constructed a biosensor containing just the ligand binding domain (LBD) of the estrogen receptor (ER).  The ER is very similar to the PPAR and other hormone receptors.  Previous attempts at isolating the LBD failed due to the specific folding pattern of this region required to maintain similar binding characteristics to natural ER.  The folding pattern was kept by inserting the LBD into a minimal splicing intein domain.  This construct was also made more soluble by addition of a maltose-binding tag.  The ER was fused with a thymidylate synthase enzyme (TS) that remains deactivated until homodimerization of the ER after binding ligand.  The cells are grown on thymine-free plates allowing for recognition of strength and function of ER ligands.</p>
 
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<p>Our plan for this project is to work on the sensitivity of the biosensor in hopes of using this for water prescreens, similar to the <em>Smart Fold Reporter</em> project. The sensitivity will be focused for BPA which has a very different conformation than the natural agonist of the ER system. This difference causes BPA to bind weakly but still disturbs normal ER function.  One idea is to replace the ER LBD from Wood’s biosensor with the estrogen-related receptor (ERR) LBD.  The ERR is similar to ER and binds many of the same ligands and has a tendency to bind to the estrogen response element (ERE) in the human body.  The one benefit of ERR for our project is that it binds BPA very strongly.  Another direction that this project could take would be to analyze the LBD of ER and perform directed evolution to increase BPA sensitivity.  During directed evolution, certain regions of the ER LBD would be targeted for random mutagenesis providing a library of mutants in the trillions.  The mutant library would be induced with BPA and the best growing colony would be selected, tested, and mutated for further sensitivity. 
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      <li><a href="http://www.princeton.edu/che/people/faculty/wood/" title="David Wood" target="_blank">David Wood</a></li>
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      <li><a href="http://www.cmtc.psu.edu/" title="Center for Molecular Toxicology and Carcinogenesis" target="_blank">PennState Center for Molecular Toxicology and Carcinogenesis</a></li>
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    <td colspan="2" style="padding-top:30px; padding-right:30px" valign="top" width="45%"><span style="font-size: 14pt">Diauxie Elimination: <em>Two</em> spoons full of sugar.</span>
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      <p><img src="picture here" alt="[img]" style="float:left; margin:5px;"/>Cellulosic biomass is an abundant and inexpensive energy source, coming from plant waste: ideal for Ethanol production through fermentation. However, biomass contains glucose and xylose sugars in relatively equal ratios, preferentially <em>e. coli</em> metabolizes glucose before any other sugar. In this project we attempt to eliminate this phenomenon, called <em>diauxie</em>, and get our cells to utilize both sugars at the same time. Solving this problem will lead to more efficent use of cellulosic biomass including moving towards the future of bioproduction continous processes.</p>
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    <h3>Drew Endy On Synthetic Biology</h3>
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   <td style="padding-top:30px; padding-right:30px" valign="top" width="45%"><span style="font-size: 14pt">Quick Links</span>
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            <p><a href="http://parts.mit.edu/igem07/index.php/PennState/final_result#contributions" title="Our Contribution To the Registry">Table of our Contributions to the Registry</a></p>
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            <p><a href="http://parts.mit.edu/igem07/images/7/73/IgemDesign.swf" title="Interactive Schematic" target="_blank">Interactive <em>E. co</em>Lisa Schematic</a></p>
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            <p> <a href="http://www.psu.edu" title="Pennsylvania State University" target="_blank">Pennsylvania State University</a> </p>
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            <p> <a href="http://www.environment.psu.edu/" title="Penn State Institutes of Energy and the Environment" target="_blank">Penn State Institutes of Energy and the Environment</a> </p>
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    <h3>Garrett Tobin On Diauxie Elimination</h3>
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Latest revision as of 03:25, 30 October 2008

Home The Team The Project Parts Notebook

Diauxie Elimination

Introduction
The System
Strategies
Progress
Conclusions
Parts
References

NHR Biosensors

NHR Introduction
Phthalate Biosensor
BPA Biosensor
 Welcome to Penn State iGEM 2008

Penn State University has participated in the International Genetically Engineered Machines (iGEM) competition for four years as of 2008, and we are excited to participate once more. Our team consists of 5 undergraduate students, one visiting undergrad and one high school student who work independently, coordinated through weekly meetings with our advisers. Check out our Team Page to meet us!

Our main focus for 2008 was the elimination of diauxie in E. coli to create a xylose inducible system independent of glucose regulation. This system could be used for creating more efficient bioproduction by altering the utilization of 5 and 6 carbon sugars. Please check out the links on the left to navigate through our work.

Two other projects begun this year focus on creating biosensors that use human nuclear hormone receptors to recognize potentially harmful compounds. These receptor systems occur naturally in the human body, but our goal is to retain and utilize their functions in E. coli. The links on the left should introduce you to our thought process and progress on each of these projects, and provide a fuller introduction to the topic.

Please explore our website to find out more about us and our projects! If there are any questions or comments about the information on this site please feel free to contact us at gjt5001@psu.edu.

Medal Checklist

Related Links

Drew Endy On Synthetic Biology

Garrett Tobin On Diauxie Elimination

Sponsors for our team! Thanks so much!

invitrogen Dupont

Retrieved from "http://2008.igem.org/Team:PennState"