Difference between revisions of "Part:BBa K3771046"
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<br>The ompA promoter facilitates the constitutive expression of OmpA/OprF. Binding of IFN-γ to the OmpA/OprF chimeric protein induces the response of the phage shock protein (Psp) system, a highly conserved stress response system in enterobacteria. [1] Signal transduction from the outer membrane to the inner membrane activates the pspA promoter, initiating expression of CDO1. CDO1 converts L-cysteine into L-cysteine sulfonic acid in the taurine synthesis L-cysteine sulfinic acid pathway [2].<br>
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<br>The <i>ompA</i> promoter facilitates the constitutive expression of OmpA/OprF. Binding of IFN-γ to the OmpA/OprF chimeric protein induces the response of the phage shock protein (Psp) system, a highly conserved stress response system in enterobacteria. [1] Signal transduction from the outer membrane to the inner membrane activates the <i>pspA</i> promoter, initiating expression of CDO1. CDO1 converts L-cysteine into L-cysteine sulfonic acid in the taurine synthesis L-cysteine sulfinic acid pathway [2].<br>
  
 
<br><b style="font-size:1.3rem">Usage</b>
 
<br><b style="font-size:1.3rem">Usage</b>
 
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<br>We ligased the NPO-OmpA* fragment and pspA-CDO1-his-tag on the pSU expression vector and transformed it into DH5alpha to complete construction of the plasmid.  The his-tag allows for confirmation of CDO1 expression by western blot using the anti-6X his-tag antibody.<br>
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<br>We ligased the <i>P<sub>ompA</sub>-ompA/oprF</i> fragment and <i>P<sub>pspA</sub>-CDO1-his-tag</i> on the pSU expression vector and transformed it into DH5α to complete construction of the plasmid.  The his-tag allows for confirmation of CDO1 expression by western blot using the anti-6X his-tag antibody.<br>
  
  

Revision as of 15:44, 17 October 2021


PpspA-CDO1-6xHis-pOmpA-OmpA/OprF


Description


This composite part is a component of the IFN-γ sensing system and was used to express the taurine production enzyme, CDO1.


Biology


The ompA promoter facilitates the constitutive expression of OmpA/OprF. Binding of IFN-γ to the OmpA/OprF chimeric protein induces the response of the phage shock protein (Psp) system, a highly conserved stress response system in enterobacteria. [1] Signal transduction from the outer membrane to the inner membrane activates the pspA promoter, initiating expression of CDO1. CDO1 converts L-cysteine into L-cysteine sulfonic acid in the taurine synthesis L-cysteine sulfinic acid pathway [2].


Usage


We ligased the PompA-ompA/oprF fragment and PpspA-CDO1-his-tag on the pSU expression vector and transformed it into DH5α to complete construction of the plasmid. The his-tag allows for confirmation of CDO1 expression by western blot using the anti-6X his-tag antibody.



Characterization


Reference


1. Darwin AJ. The phage-shock-protein response. Molecular Microbiology. 2005;57(3):621-628. doi:10.1111/j.1365-2958.2005.04694.x https://pubmed.ncbi.nlm.nih.gov/16045608/
2. Joo Y-C, Ko YJ, You SK, et al. Creating a New Pathway in Corynebacterium glutamicum for the Production of Taurine as a Food Additive. Journal of Agricultural and Food Chemistry. 2018;66(51):13454-13463. doi:10.1021/acs.jafc.8b05093 https://pubmed.ncbi.nlm.nih.gov/30516051/


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 12
    Illegal BamHI site found at 1915
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 591
  • 1000
    COMPATIBLE WITH RFC[1000]