Difference between revisions of "Team:Vilnius-Lithuania/Demonstrate"

 
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<img src="https://static.igem.org/mediawiki/2018/6/6e/T--Vilnius-Lithuania--Simo_fig6.jpg">
 
<img src="https://static.igem.org/mediawiki/2018/6/6e/T--Vilnius-Lithuania--Simo_fig6.jpg">
<P> Proteins that possess ß-barrel structures do not fully denature in the presence of SDS if they are unboiled, which leads to unorthodox movement in SDS-PAGE. As BamA only forms ß-barrels when correctly folded, we can evaluate the amount of protein correctly folded and incorporated into the liposome membranes by the amount of unusual product in an SDS-PAGE.
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<p> Proteins that possess ß-barrel structures do not fully denature in the presence of SDS if they are unboiled, which leads to unorthodox movement in SDS-PAGE. As BamA only forms ß-barrels when correctly folded, we can evaluate the amount of protein correctly folded and incorporated into the liposome membranes by the amount of unusual product in an SDS-PAGE.
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<p><a href="https://2018.igem.org/Team:Vilnius-Lithuania/Design#BAM%20complex">More about Bam complex proteins</a>
 
<p><a href="https://2018.igem.org/Team:Vilnius-Lithuania/Design#BAM%20complex">More about Bam complex proteins</a>
 
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             <li>Finally, as our ultimate goal was to use liposomes as a platform for membrane protein research, we constructed membrane proteins that were capable of displaying protein particles on the surface of E. coli, hence showed no warning signs of not being to do the same in liposomes.</li>>
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             <li>Finally, as our ultimate goal was to use liposomes as a platform for membrane protein research, we constructed membrane proteins that were capable of displaying protein particles on the surface of E. coli, hence showed no warning signs of not being to do the same in liposomes.</li>
 
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<p>We successfully constructed multiple novel membrane proteins based on ß-barrel bearing translocators that are capable of displaying protein particles on the surface of E. coli (Fig. 6). As liposomes feature the same lipids as E. coli membranes, and during our project were successfully incorporated with Bam complex proteins, it appears that these proteins will be able to expose fused protein particles as, if not more, efficiently as in bacteria, laying the foundation to reach new molecular evolution horizons.
 
<p>We successfully constructed multiple novel membrane proteins based on ß-barrel bearing translocators that are capable of displaying protein particles on the surface of E. coli (Fig. 6). As liposomes feature the same lipids as E. coli membranes, and during our project were successfully incorporated with Bam complex proteins, it appears that these proteins will be able to expose fused protein particles as, if not more, efficiently as in bacteria, laying the foundation to reach new molecular evolution horizons.
 
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<img src="https://static.igem.org/mediawiki/2018/4/47/T--Vilnius-Lithuania--Simui_bac_expo.jpg">
 
<img src="https://static.igem.org/mediawiki/2018/4/47/T--Vilnius-Lithuania--Simui_bac_expo.jpg">
<strong>Fig. 7</strong> Plate reader results of the absorption at 450 nm wavelength of the bacteria coding for different exposition proteins. Absorption’s intensity is proportional to the efficiency of display.
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<p><strong>Fig. 7</strong> Plate reader results of the absorption at 450 nm wavelength of the bacteria coding for different exposition proteins. Absorption’s intensity is proportional to the efficiency of display.</p>
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<p><a href="https://2018.igem.org/Team:Vilnius-Lithuania/Design#Surface_display_system">More about surface display</a>
 
<p><a href="https://2018.igem.org/Team:Vilnius-Lithuania/Design#Surface_display_system">More about surface display</a>

Latest revision as of 21:28, 4 November 2018

Proof of Concept

The Composite Proof

We proved that our SynDrop system worked as intended by successfully implementing several critical wet lab and dry lab experiments. First, we have created a model to determine microfluidics variables for optimal liposome synthesis. Second, we synthesized stable biocompatible liposomes and demonstrated an internal transcription and translation of functional proteins. Third, we demonstrated that membrane proteins can successfully integrate into our liposomes. Finally, we constructed working fusion proteins that were able to display a designated tag on the outer membrane.

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