Team:UC Davis/Project1

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<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a
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href="https://2009.igem.org/Team:UC_Davis/homepage2"><img alt=""
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style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a
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href="https://2009.igem.org/Team:UC_Davis/About_Us1"><b
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a
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style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a
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href="file:///C:/Davis/iGEM09/igem2009_site/UCDAVIS_Contacts.html"><img
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</b>
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<hr style="width: 100%; height: 2px;">
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<div style="text-align: left;"><b style="color: rgb(0, 0, 0);"><big><big>Project:</big></big><br>
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</b>
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<div style="text-align: center;"><b style="color: rgb(0, 0, 0);"><a
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href="#celiac_disease">Celiac
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Disease</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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<a href="#current_treatment">Current
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treatment</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<a
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href="#our_approach">
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Our Approach</a></b><br>
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</div>
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<small><u><span
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style="font-size: 20pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span
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style="font-weight: bold;"><a name="celiac_disease"></a>Celiac Disease:</span><br>
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</span></u></small><i style="font-weight: bold;"><span
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style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br>
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</span></i><i><span
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style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">What
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is Celiac Disease?</span></i><br>
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<i><u><span
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style="font-size: 20pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></u></i><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;
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&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Celiac
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Disease is a form of autoimmune disorder, which occurs inside the small
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intestine. When the body cannot digest gliadin (component of gluten)
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properly,
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this leads to an immune response (14). Furthermore, different people
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have
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different reactions to these immune responds such as pain and vomiting
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(14). It
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has been estimated that about one out of 133 Americans are currently
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suffering
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from this disease (11); including one of our teammate's friends, and
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that was
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what sparked our interest in designing this project.</span><span
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style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</span></span><span
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style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span
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style=""></span><span style=""></span></span><span
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style="font-style: italic;"><span
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style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br>
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<br>
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What happens in the normal small
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intestine?</span></span><span
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style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br>
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&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; In
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the normal small intestine, the Brush border membrane lets small
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peptides and
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molecules like water through and into the bloodstream. The brush border
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membrane also lets larger molecules into the blood stream, similar to
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Gliadin;
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however, these larger molecules are converted through a transcellular
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route, which
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does not prompt an immune system response.<o:p></o:p></span><i><span
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style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br>
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<br>
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What
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happens in celiac disease small intestine?</span></i><i><span
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style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span
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style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</span><o:p></o:p></span></i><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br>
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&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; The
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Brush border membrane in the small intestine of people suffering from
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Celiac Disease
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allows large molecules such as gliadin to go straight into the blood
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stream
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without directing them through the transcellular route. Once gliadin
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passes
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through, Antigen Presenting Cells (APCs) recognize gliadin as a foreign
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object
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and will attack. This immune response causes pain and other adverse
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side-effects.<o:p></o:p></span>
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<p class="MsoNormal" style="text-indent: 0.5in;"><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">The
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immune response not only causes abdominal bloating, pain, weight loss
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and
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vomiting, most of the time it may also damage intestinal villi, which
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are
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important for absorbing nutrition. The destruction of villi eventually
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leads to
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a lack of absorption of different nutrients, which can lead to other
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illnesses
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(14). Some people affected with this disorder can also suffer from </span><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">autoimmune
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thyroid disease, autoimmune liver disease, rheumatoid arthritis </span><i><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">(diseases
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in which body immune system attacks healthy cells/tissues)</span></i><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">
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(14).</span><b><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br>
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</span></b></p>
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<hr style="width: 100%; height: 2px;">
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<p class="MsoListParagraph" style=""><u><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><span
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style="font-weight: bold;"><a name="current_treatment"></a>Current
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treatments:</span></big></big><o:p></o:p></span></u></p>
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<p class="MsoNormal" style="line-height: normal;"><b><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</span></b><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Currently,
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no cure has been found
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for this illness. The only way to avoid some of celiac disease’s
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symptoms is by
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adopting a gluten free diet (14, 12, and 11). However, you are more
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likely to
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find gluten in your everyday diet. Therefore, avoiding gluten is not
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easy or
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pleasant from a gastronomical standpoint.<br>
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</span><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p>
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&nbsp; &nbsp; &nbsp; &nbsp; &nbsp;&nbsp;
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"Oral supplementation with prolyl oligopeptidases has therefore been
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proposed
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as a potential therapeutic approach."(8). However, enzymes studied
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earlier
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were not able to degrade gluten (inside stomach before it reaches small
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intestine because they were "irreversibly inactivated by pepsin and
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acidic
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pH, both present in the stomach."(8)<o:p></o:p></span><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br>
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&nbsp; &nbsp; &nbsp;&nbsp; &nbsp;&nbsp;&nbsp;&nbsp;
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Nevertheless, over the past years, researchers have discovered an
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enzyme from <i>Aspergillus
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niger</i>, a newly identified prolyl endoprotease, that was observed to
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"work optimally at 4-5pH and remains stable at 2pH"(8) and maybe this
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enzyme will lead us to an alternative treatment for this disorder (8).
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Studies
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have shown that prolyl-endoprotease from <i>Aspergillus niger</i> is
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able to
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"degrade gluten in vitro and under conditions similar to the ones
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present
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in the gastrointestinal tract." (8) ; but due to licensing restrictions
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we
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have opted not to work with this protein.</span><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span><br>
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<span style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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Recently (year 2007), a study has suggested an alternative approach by
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combining a glutamine-specific endoprotease (EP-B2 from barley) and a
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prolyl
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endopeptidase (SC PEP from Sphingomonas capsulata); with gastric
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activity and
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complementary substrate specificity there is a possibility of
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increasing the
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safe threshold of ingested gluten (12). One of the advantages of this
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“combination product is that both enzymes are active and stable in
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stomach and
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can therefore be administered as lyophilized powders or simple capsules
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or
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tablets” (12).</span><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p>
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<p class="MsoNormal" style="line-height: normal;"><i><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Note:
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Moreover, this study was first
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“evaluated via in vitro digestion of whole-wheat beard and then
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confirmed by in
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vivo studies in rats (12), unlike other earlier studies which were
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performed on
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synthetic gluten oligopeptides, recombinant gliadin proteins, or
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uncooked
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gluten” (12).</span></i><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p>
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<hr style="width: 100%; height: 2px;"><b><u><span
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style="font-size: 20pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><a
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name="our_approach"></a>Our
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approach:<o:p></o:p></span></u></b>
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<p class="MsoNormal"><span
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style="font-size: 16pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span
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style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span></span><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">We
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are going to break down gluten inside the stomach (before it arrives to
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the
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small intestine), because gliadin does not prompt an immune system
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response until
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it is absorbed in the small intestine.<span style="">&nbsp;
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</span>Another worry is a spread of our synthetic bug, so we engineered
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a
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‘safety-switch’ via induction of cell death through a pH sensor system.<span
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style="">&nbsp; </span>Due to the change of pH from the stomach to
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the small intestine, our pH system is induced; cell death, thus
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limiting our
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bug to the stomach.<o:p></o:p></span></p>
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<p class="MsoNormal"><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span
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style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</span>The
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advantages of our model are, that unlike other suggested treatments
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which
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require consuming capsules or tablets rapidly, by using <i><span
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style="color: black;">Escherichia coli</span></i><span
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style="color: black;"> as
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our model chassis for this project: we can lower the number of times
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which this
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‘cure’ needs to be consumed thus making it less troublesome, costly,
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and more
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convenient.<br>
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</span></span></p>
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<hr style="width: 100%; height: 2px;">
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<p class="MsoNormal"><span style="font-weight: bold;">Our project is
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split into two parts:</span><br>
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1.Induced Secretion<br>
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</p>
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<ul>
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<li>Gene sequence for secretion system
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</li>
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<li>Adding Secretion
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</li>
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<li>Assay for measuring protein localization and activity</li>
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</ul>
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2.Sensing pH&nbsp; and inducing cell death<br>
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<ul>
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<li>Finding an appropriate biological pH sensor
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</li>
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<li>Wiring the pH sensor into our system
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</li>
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<li>Assay for measuring the behavior of the pH sensor
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</li>
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</ul>
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<hr style="width: 100%; height: 2px;">
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<p class="MsoNormal"><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span
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style="color: black;"></span></span><b><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span
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style="color: black;"><o:p></o:p></span></span></b></p>
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</div>
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style="font-size: 18pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><small></small>
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</span></small><!--[endif]--><span
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</div>
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Latest revision as of 21:40, 25 September 2009

Retrieved from "http://2009.igem.org/Team:UC_Davis/Project1"