Difference between revisions of "Team:Bielefeld-CeBiTec/Results/toolbox"

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<h2> Short summary </h2>
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Our aim was to provide an innovative toolbox to incorporate different noncanonical amino acids (ncAAs) during translation to expand the possibilities in protein design. We choose seven different ncAAs in our five tools and demonstrate them all in interesting applications. These ncAAs can be used for a lot of different approaches in basic research, medicine and manufacturing and with our submitted parts, every iGEM team can incorporate these seven ncAAs into their target proteins. A short description of every tool is listed below.
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<h3> Analysing </h3>
 
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The structural analysis of a protein can be used to study distances between noncanonical amino acids (ncAAs) with Foerster Resonance Energy Transfer (FRET). This provides measuring distances between specific incorporated amino acids in the target protein. To incorporate the ncAAs we provide three different tRNA/aminoacyl-synthetases which incorporate in respose to the amber or the less used leucine codon. To demonstrate this tool we also developed a prion detection assay.
 
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<h3> Labeling </h3>
 
<h3> Labeling </h3>
 
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The labeling of a protein in vivo is a useful tool that allows the investigation of a protein in its native environment. As a label for our target protein we use the fluorescent amino acid L-(7 hydroxycoumarin 4 yl) ethylglycine (CouAA) that is incorporated by an orthogonal t-RNA/aminoacyl synthethase pair (tRNA/aaRS) at a defined position. We provide this tRNA/aaRS to the iGEM community and demonstrated this tool with the colocalization of the artificial cell compartment carboxysome and the ribulose 1,5-bisphosphate carboxylase oxygenase.
 
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Revision as of 12:16, 12 October 2017

Toolbox Overview

Short summary

Our aim was to provide an innovative toolbox to incorporate different noncanonical amino acids (ncAAs) during translation to expand the possibilities in protein design. We choose seven different ncAAs in our five tools and demonstrate them all in interesting applications. These ncAAs can be used for a lot of different approaches in basic research, medicine and manufacturing and with our submitted parts, every iGEM team can incorporate these seven ncAAs into their target proteins. A short description of every tool is listed below.

Analysing

The structural analysis of a protein can be used to study distances between noncanonical amino acids (ncAAs) with Foerster Resonance Energy Transfer (FRET). This provides measuring distances between specific incorporated amino acids in the target protein. To incorporate the ncAAs we provide three different tRNA/aminoacyl-synthetases which incorporate in respose to the amber or the less used leucine codon. To demonstrate this tool we also developed a prion detection assay.

Photoswichting

We designed the concept of a light influenced lycopene production as basis for the establishment of a photo switchable enzyme activity. We introduced an amber-codon in the gene for crtI in the lycopene pathway and thus stopped the lycopene production. We then showed that the lycopene production can be partially retained, by cotransforming with an aaRS and cultivation without the matching ncAA.

Labeling

The labeling of a protein in vivo is a useful tool that allows the investigation of a protein in its native environment. As a label for our target protein we use the fluorescent amino acid L-(7 hydroxycoumarin 4 yl) ethylglycine (CouAA) that is incorporated by an orthogonal t-RNA/aminoacyl synthethase pair (tRNA/aaRS) at a defined position. We provide this tRNA/aaRS to the iGEM community and demonstrated this tool with the colocalization of the artificial cell compartment carboxysome and the ribulose 1,5-bisphosphate carboxylase oxygenase.