Expression, Purification and structural characterization of Escherichia coli OxyR protein as a peroxide sensor

OxyR is a peroxide-sensitive and transcription regulator of the LysR-type family in bacteria. In the presence of hydrogen peroxide, a subset of antioxidant genes is expressed and the cell is protected against oxidative stress conditions. H2O2 changes the structure of OxyR in the regulatory region of the protein via a di-sulfide bond formation between the cysteines, and therefore, induces the activity of antioxidant enzymes. In this study, by investigating the catalase activity of isolated strains, the best strain as the best antioxidant was selected. Then, the nucleotide sequence of the OxyR gene was subcloned into the prokaryotic expression vector pET-21a, and induced by IPTG in E. coli BL21 expression system. In the following, OxyR protein was purified by Ni-NTA chromatography. The oxyR purity was determined using 12% SDS-PAGE and oxyR structure was studied by FTIR and NMR. The results revealed the high purity and tetrameric assembly of isolated oxyR. These findings may help to expand the understanding of redox-related proteins purification strategies.

کلیدواژه ها:

Protein structure ، Antioxidant response ، E-Coli ، OxyR protein