Difference between revisions of "Part:BBa K3031018"

Revision as of 16:10, 21 October 2019

OxyR mutated at residues 147 & 203

OxyR Transcription factor protein. OxyR reacts with H2O2 resulting in the oxidation of a reactive cysteine (Cys-199) formation of a intermolecular disulfide bond and thus a conformation change in the OxyR protein. The activated OxyR then binds to certain promoter regions to regulate the expression of downstream genes.

This is an improvement to the part BBa_K1104200 submitted by National Yang Ming University iGEM Team 2013.

Contribution: Improvement and Characterization

Team SUIS_Shanghai 2019 attempted to improve the sensitivity of this OxyR to reactive oxygen species (ROS) by increasing the accessible surface area of the reactive cysteine (cysteine-199) within the protein. This cysteine, once oxidized will form a intermolecular disulfide bond with Cys-208, resulting in a conformational change which can activate certain ROS sensitive promoters and therefore positively expressing genes in times of oxidative stress. Our approach was to use a large set of statistical data to identify certain structural features within the environment of reactive cysteines in a wide variety of protein structures sourced from the Protein Data Bank (PDB). We compared the structural features of these oxidized cysteine environments (the cysteines will form a Sulfenic acid upon oxidation – a post translational modification which is known to be a switch in some proteins) with unmodified cysteine residues within the same proteins. The information we obtained from this large study (of over 300 different proteins) was then used to inform decisions into editing the OxyR gene sequence to mutate the amino acid sequence around the Cys-199. We hoped to make this transcription factor more sensitive to lower concentrations of H2O2 and potentially create a new device which could be used to be a very sensitive indicator of early immune response and infection.

To obtain a comprehensive account of the data used to make the decisions to edit the OxyR nucleotide sequence, please visit our Model page. Our analysis led us to change two amino acids in the OxyR protein surrounding the Cys-199

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Sequence and Features