Team:Uppsala/signal-peptide

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                                                 <li><a href="https://2013.igem.org/Team:Uppsala/metabolic-engineering">Metabolic engineering</a>
                                                 <li><a href="https://2013.igem.org/Team:Uppsala/metabolic-engineering">Metabolic engineering</a>
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                                                                 <li><a href="https://2013.igem.org/Team:Uppsala/p-coumaric-acid">p-Coumaric acid</a></li>
                                                                 <li><a href="https://2013.igem.org/Team:Uppsala/resveratrol">Resveratrol</a></li>
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<li><a href="https://2013.igem.org/Team:Uppsala/lycopene">Lycopene</a></li>
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<li><a href="https://2013.igem.org/Team:Uppsala/chromoproteins">Chromoproteins</a></li>
<li><a href="https://2013.igem.org/Team:Uppsala/chromoproteins">Chromoproteins</a></li>
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<li><a href="https://2013.igem.org/Team:Uppsala/P-Coumaric-acid-pathway">P-Coumaric acid</a></li>
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<li><a href="https://2013.igem.org/Team:Uppsala/P-Coumaric-acid-pathway">Kinetic model</a></li>
<li><a href="https://2013.igem.org/Team:Uppsala/modeling-tutorial">Modeling tutorial </a></li>
<li><a href="https://2013.igem.org/Team:Uppsala/modeling-tutorial">Modeling tutorial </a></li>
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<li><a href="https://2013.igem.org/Team:Uppsala/carotenoid-group">Carotenoid group</a></li>
<li><a href="https://2013.igem.org/Team:Uppsala/carotenoid-group">Carotenoid group</a></li>
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<li><a href="https://2013.igem.org/Team:Uppsala/bioart">BioArt</a></li>
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<h1> How signal peptides work </h1>
<h1> How signal peptides work </h1>
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<p>A signal peptide, also called leader peptide, is the first part of extracellular proteins and can vary in length, its a short sequence fused to the N-terminus of a protein. The ones we work with are 25 amino acids and 50 amino acids in length. The signal peptides enables the protein to be translocated through the bacterial plasma membrane via the SecY complex.<sup><a href="#l1">[1]</a></sup></p>   
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<p>A signal peptide, also called leader peptide, is the first part of extracellular proteins and can vary in length, its a short sequence fused to the N-terminus of a protein. The signal peptides we work with are 25 and 50 amino acids in length. The signal peptides enables the protein to be translocated through the bacterial plasma membrane via the SecY complex.<sup><a href="#l1">[1]</a></sup></p>   
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<a id="b1"></a>
<h1> Lactobacillus - A champion of protein secretion </h1>
<h1> Lactobacillus - A champion of protein secretion </h1>
<p>One of the specific advantageous traits of the lactobacillus genus is that they are great at secreting proteins. Therefore we decided to synthesize the signal peptide usp45 from lactococcus lactis according to the Freiburg fusion standard to enable fusion with any protein. This peptide has been shown to work in lactobacillus reuteri together with GFP and will probably work in other lactobacillus species.<sup><a href="#l2">[2]</a></sup></p>
<p>One of the specific advantageous traits of the lactobacillus genus is that they are great at secreting proteins. Therefore we decided to synthesize the signal peptide usp45 from lactococcus lactis according to the Freiburg fusion standard to enable fusion with any protein. This peptide has been shown to work in lactobacillus reuteri together with GFP and will probably work in other lactobacillus species.<sup><a href="#l2">[2]</a></sup></p>
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<img class="method-plasmid" src="https://static.igem.org/mediawiki/2013/9/9d/Uppsala2013_signal-peptides.jpg">
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<a href="https://static.igem.org/mediawiki/2013/9/9d/Uppsala2013_signal-peptides.jpg" data-lightbox="roadtrip"><img class="method-plasmid" src="https://static.igem.org/mediawiki/2013/9/9d/Uppsala2013_signal-peptides.jpg"></a>
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<p>One of the goals in our project was to make a probiotic bacteria express the protein miraculin (Mir in picture) which would work as a sweetener. However because it works by binding to receptors on the tongue and the protein is too big to pass through the cell membrane of the bacteria it needs to be actively secreted to work optimally. Fusing a signal peptide to the N-terminus of miraculin can solve this. We have synthesised miraculin fused together with usp45. We have not yet had the chance to transform and characterise it in Lactobacillus.</p>
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<p>One of the goals in our project was to make a probiotic bacteria express the protein miraculin (Mir in picture) which would work as a sweetener. However because it works by binding to receptors on the tongue and the protein is too big to pass through the cell membrane of the bacteria it needs to be actively secreted to work optimally. Fusing a signal peptide to the N-terminus of miraculin can solve this. We have synthesised miraculin fused together with usp45. Because we received the synthesised miraculin gene late, we did not have time to put it in pSB1C3 and send it to the registry. We have also not yet had the chance to transform and characterise it in Lactobacillus.</p>
<h1> References: </h1>  
<h1> References: </h1>  
<a name="l1">[1]</a> Rapoport T. (Nov. 2007). "Protein translocation across the eukaryotic endoplasmic reticulum and bacterial plasma membranes.". Nature 450 (7170): 663–9.
<a name="l1">[1]</a> Rapoport T. (Nov. 2007). "Protein translocation across the eukaryotic endoplasmic reticulum and bacterial plasma membranes.". Nature 450 (7170): 663–9.
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<br><br>
<a name="l2">[2]</a> Chi-Ming Wu. (April 2006). “Green fluorescent protein is a reliable reporter for screening signal peptides functional in Lactobacillus reuteri” Journal of Microbiological Methods (2006) 181-186.
<a name="l2">[2]</a> Chi-Ming Wu. (April 2006). “Green fluorescent protein is a reliable reporter for screening signal peptides functional in Lactobacillus reuteri” Journal of Microbiological Methods (2006) 181-186.
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Latest revision as of 21:30, 28 October 2013

Signal peptides

How signal peptides work

A signal peptide, also called leader peptide, is the first part of extracellular proteins and can vary in length, its a short sequence fused to the N-terminus of a protein. The signal peptides we work with are 25 and 50 amino acids in length. The signal peptides enables the protein to be translocated through the bacterial plasma membrane via the SecY complex.[1]

Lactobacillus - A champion of protein secretion

One of the specific advantageous traits of the lactobacillus genus is that they are great at secreting proteins. Therefore we decided to synthesize the signal peptide usp45 from lactococcus lactis according to the Freiburg fusion standard to enable fusion with any protein. This peptide has been shown to work in lactobacillus reuteri together with GFP and will probably work in other lactobacillus species.[2]

One of the goals in our project was to make a probiotic bacteria express the protein miraculin (Mir in picture) which would work as a sweetener. However because it works by binding to receptors on the tongue and the protein is too big to pass through the cell membrane of the bacteria it needs to be actively secreted to work optimally. Fusing a signal peptide to the N-terminus of miraculin can solve this. We have synthesised miraculin fused together with usp45. Because we received the synthesised miraculin gene late, we did not have time to put it in pSB1C3 and send it to the registry. We have also not yet had the chance to transform and characterise it in Lactobacillus.

References:

[1] Rapoport T. (Nov. 2007). "Protein translocation across the eukaryotic endoplasmic reticulum and bacterial plasma membranes.". Nature 450 (7170): 663–9.

[2] Chi-Ming Wu. (April 2006). “Green fluorescent protein is a reliable reporter for screening signal peptides functional in Lactobacillus reuteri” Journal of Microbiological Methods (2006) 181-186.