Team:IIT Delhi 1

From 2010.igem.org

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<div style='mso-element:para-border-div;border:none;border-bottom:solid #4F81BD 1.0pt;
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<p align=center>[[Image:IGEM_IITD_logo.png|200px]]</p>
 
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<p class=MsoTitle align=center style='text-align:center'><span lang=EN-US><i style='mso-bidi-font-style:
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<p align=center><embed src="https://static.igem.org/mediawiki/igem.org/5/51/Nish1.swf" width=900 height=500></p>
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normal'><b>Dr. Coli </b></i></span></p>
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</div>
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<p class=MsoNormal><span lang=EN-US></span></p>
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<h2><span lang=EN-US><u>Renin Angiotensin Aldosterone System (RAAS)<o:p></o:p></span></u></h2>
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<p class=MsoNormal><span lang=EN-US><o:p>&nbsp;</o:p></span></p>
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<h3><span lang=EN-US>Introduction</span></h3>
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<p class=MsoNormal style='text-align:justify'><span lang=EN-US>The Renin
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Angiotensin Aldosterone System is a hormone system in the human body that is
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responsible for the regulation of blood pressure and fluid balance.</span></p>
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<h3 style='text-align:justify'><span lang=EN-US>Functioning</span></h3>
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<p class=MsoNormal style='text-align:justify'><span lang=EN-US>A decrease in
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renal perfusion causes the release of the enzyme renin which subsequently
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cleaves angiotensinogen to angiotensin I. This is further converted to
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angiotensin II by angiotensin-converting enzyme (ACE). The bioactive product,
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angiotensin II, causes the constriction of blood vessels, leading to an
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increased blood pressure. It is also responsible for the secretion of
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aldosterone by the adrenal cortex, which acts by increasing the re-absorption
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of sodium and water into the blood. Hence, the fluid volume as well as the
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blood pressure is maintained in the body.</span></p>
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<h2><u><span lang=EN-US>The &#945;-hemolysin System in <i style='mso-bidi-font-style:
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normal'>E.coli<o:p></o:p></i></span></u></h2>
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<p class=MsoNormal><span lang=EN-US><o:p>&nbsp;</o:p></span></p>
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<h3><span lang=EN-US>Introduction</span></h3>
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<p class=MsoNormal style='text-align:justify'><span lang=EN-US>The
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alpha-hemolysin system is a type I secretion system found in wild type <i
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style='mso-bidi-font-style:normal'>E.coli </i>which enable them to cause
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infection in the urinary tract and is an important virulence factor because of
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its cytotoxic and cytolytic activity against a variety of mammalian cells. </span></p>
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<h3 style='text-align:justify'><span lang=EN-US>Functioning</span></h3>
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<p class=MsoNormal style='text-align:justify'><span lang=EN-US>The hemolysin
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system originates from a series of hly genes. These genes are present in the
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series <i style='mso-bidi-font-style:normal'>hlyC – hlyA – hlyB – hlyD</i>. Of
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these, <i style='mso-bidi-font-style:normal'>hlyA</i> codes for the protein
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that is responsible for the infection. Hly B and D are membrane proteins that
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work in coordination for the secretion of HlyA. The synthesis, activation and
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secretion of HlyA in <i style='mso-bidi-font-style:normal'>E.coli </i>is
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determined by the <i style='mso-bidi-font-style:normal'>hlyCABD</i> operon. This
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operon is either located either on the chromosome or on transmissible plasmids.
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</span></p>
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<p class=MsoNormal style='text-align:justify'><span lang=EN-US>It has been
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shown that this system can be used to produce and secrete a protein of our
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choice if the protein contains 60 amino acid residues from the carboxy terminal
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end of the HlyA protein. </span></p>
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<h2><u><span lang=EN-US>Our iGEM 2010 Project<o:p></o:p></span></u></h2>
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<p class=MsoNormal><span lang=EN-US><o:p>&nbsp;</o:p></span></p>
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<h3><span lang=EN-US>Introduction</span></h3>
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<p class=MsoNormal style='text-align:justify'><span lang=EN-US>Our aim is to
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create an online elicitor detection and response in a flow system, which can
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function in the detection of high blood pressure and help reduce it by
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releasing renin inhibitors into the circulatory system. This kind of a system
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would be helpful to patients with a hypertensive condition. </span></p>
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<h3><span lang=EN-US>Approach</span></h3>
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<p class=MsoNormal><span lang=EN-US>Since the blood pressure system is not
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really a chemical signal, what we have assumed is that the blood pressure can
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be easily converted into a chemical signal. Having assumed this, we will use a
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chemical signal such as IPTG or AHL as the elicitor to produce appropriate
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amounts of renin inhibitor. For easy detection and verification, we are
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planning to use GFP in place of renin inhibitor which can easily be replaced
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later.</span></p>
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<p class=MsoNormal><span lang=EN-US>For this we will be introducing two vectors
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into <i style='mso-bidi-font-style:normal'>E. coli </i>as shown below.</span></p>
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<p class=MsoNormal><span style='mso-ansi-language:EN-IN;mso-fareast-language:
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EN-IN;mso-bidi-language:HI;mso-no-proof:yes'><!--[if gte vml 1]><v:shapetype
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<v:stroke joinstyle="miter"/>
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alt="Vector1.png" style='width:195pt;height:209.25pt;visibility:visible;
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mso-wrap-style:square'>
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<v:imagedata src="IIT+Delhi+iGEM+Wiki_files/image001.png" o:title="Vector1"/>
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</v:shape><![endif]--><![if !vml]><img width=260 height=279
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src="IIT+Delhi+iGEM+Wiki_files/image002.gif" alt=Vector1.png v:shapes="Picture_x0020_0"><![endif]></span><span
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lang=EN-US><span style='mso-spacerun:yes'>              </span><span
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style='mso-tab-count:1'>          </span></span><span style='mso-ansi-language:
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EN-IN;mso-fareast-language:EN-IN;mso-bidi-language:HI;mso-no-proof:yes'><!--[if gte vml 1]><v:shape
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style='width:195pt;height:212.25pt;visibility:visible;mso-wrap-style:square'>
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<v:imagedata src="IIT+Delhi+iGEM+Wiki_files/image003.png" o:title="Vector2"/>
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</v:shape><![endif]--><![if !vml]><img width=260 height=283
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src="IIT+Delhi+iGEM+Wiki_files/image004.gif" alt=Vector2.png v:shapes="Picture_x0020_1"><![endif]></span></p>
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<p class=MsoNormal><span lang=EN-US><o:p>&nbsp;</o:p></span></p>
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<p class=MsoNormal style='text-align:justify'><span lang=EN-US>The first vector
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which contains the GFP-hlyA chimeric sequence with pLac promoter is used to
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produce the chimera GFP protein which contains 62 amino acid residues from the
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C terminal of hlyA protein, a prerequisite for the secretion of the protein
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outside the cell body. Forming a chimera will not in any way change the
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structure or function as per literature.</span></p>
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<p class=MsoNormal><span lang=EN-US><o:p>&nbsp;</o:p></span></p>
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<p class=MsoNormal><span lang=EN-US>The other vector contains the hlyB and hlyD
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sequences with ptet promoter sequence. These two proteins will be
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constitutively expressed. These proteins are present in the membrane and
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provide for an outlet of hlyA or chimeras with 62 amino acid residues of the C
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terminal of hlyA.</span></p>
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<p class=MsoNormal><span lang=EN-US>After we are successful in obtaining
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GFP-hlyA chimera outside the cell body, our next task will be to immobilize
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these cells in a flow system and characterize the flow-expression profile. We
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would want to keep the cells in G0 phase so as to have continuous expression of
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the protein without cell death</span></p>
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<p class=MsoNormal><span lang=EN-US>We will also be working towards minimizing
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the toxins and other products so that such a system can finally be thought fit
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for incorporating and integrating with the circulatory system in our body.</span></p>
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<p class=MsoNormal><span lang=EN-US><o:p>&nbsp;</o:p></span></p>
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<p class=MsoNormal style='text-align:justify'><span lang=EN-US><o:p>&nbsp;</o:p></span></p>
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<p><h2>Special comments:</h2>
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1.Plasmids acquired from Karolinska Institutet, Solna, Sweden. <br>
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2.Safety page in 'Work' link.
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Latest revision as of 01:05, 28 October 2010

Special comments:

1.Plasmids acquired from Karolinska Institutet, Solna, Sweden.
2.Safety page in 'Work' link.

Retrieved from "http://2010.igem.org/Team:IIT_Delhi_1"