Difference between revisions of "Part:BBa K3846364"

 
 
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Since CYP P450 enzymes always need their reductase aim our project was to fuse CYPs with reductase to increase reactivity of the enzymes. The effect of fusing CYPs with reductase needed to be investigated by comparing fusion proteins with unmodified parallel expressed enzymes. For that reason we designed control vectors consisting of expression cassettes CYP/reductase pairs.
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These expression cassettes contained an IPTG inducible T7 promoter, the coding sequence and a double terminator and can be combined with each other.
  
  
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<partinfo>BBa_K3846364 parameters</partinfo>
 
<partinfo>BBa_K3846364 parameters</partinfo>
 
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===iGEM Hamburg 2021 part collection===
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Terpenoids are an important group of natural products used as biofuels, drugs or fragrances.  Naturally occuring in plants it has been shown that microbial terpene production in microorganisms like yeast, E. coli or  cyanobacteria is possible.
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Nevertheless iGEM projects seem to rarely focus on this interesting class of natural products which is correlated with a lack of useful parts inside the iGEM registry.
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Fortunately we were able to change that and designed a novel golden gate based toolbox which allows.
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<ol style="list-style-type:lower-alpha">
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  <li>production of terpenoid precursors and simple terpenoids</li>
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  <li>creation of CYP P450-reductase fusion enzymes to optimise processing of terpenoid precursors and production of bioactive target products</li>
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  <li>modularity of the system to enable exchange of linker sequences/promoters/etc. (MoClo-compatible toolbox)</li>
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</ol>
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===MoClo-based Part Design 2.0===
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<p>To improve the usefulness of our parts, we then aimed to make our parts compatible with the MoClo standard of goten gate based IIS restriction enzyme assembly. Thereby we expanded the Common Genetic Syntax for fusion sites to allow the creation of a) fusion proteins connected by linker sequences and b) multiple CDS expressed in an operon.
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More useful information and an overview of all our parts can be found on our [https://2021.igem.org/Team:Hamburg/Part_Collection wiki].
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[[File:T--Hamburg--parts overview MolClo.png|600px|thumb|left|'''Figure 1''': <b> MoClo syntax of the part collection. </b> <br>]]

Latest revision as of 12:52, 20 October 2021


Ferredoxin expression cassette (T7 promoter)


Since CYP P450 enzymes always need their reductase aim our project was to fuse CYPs with reductase to increase reactivity of the enzymes. The effect of fusing CYPs with reductase needed to be investigated by comparing fusion proteins with unmodified parallel expressed enzymes. For that reason we designed control vectors consisting of expression cassettes CYP/reductase pairs. These expression cassettes contained an IPTG inducible T7 promoter, the coding sequence and a double terminator and can be combined with each other.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]



iGEM Hamburg 2021 part collection

Terpenoids are an important group of natural products used as biofuels, drugs or fragrances. Naturally occuring in plants it has been shown that microbial terpene production in microorganisms like yeast, E. coli or cyanobacteria is possible. Nevertheless iGEM projects seem to rarely focus on this interesting class of natural products which is correlated with a lack of useful parts inside the iGEM registry.

Fortunately we were able to change that and designed a novel golden gate based toolbox which allows.

  1. production of terpenoid precursors and simple terpenoids
  2. creation of CYP P450-reductase fusion enzymes to optimise processing of terpenoid precursors and production of bioactive target products
  3. modularity of the system to enable exchange of linker sequences/promoters/etc. (MoClo-compatible toolbox)

MoClo-based Part Design 2.0

To improve the usefulness of our parts, we then aimed to make our parts compatible with the MoClo standard of goten gate based IIS restriction enzyme assembly. Thereby we expanded the Common Genetic Syntax for fusion sites to allow the creation of a) fusion proteins connected by linker sequences and b) multiple CDS expressed in an operon. More useful information and an overview of all our parts can be found on our wiki.

Figure 1: MoClo syntax of the part collection.