Structure and conformational studies on dityrosine formation in the DNA binding domain of RFX5.
The DNA binding protein RFX5 is a subunit of RFX complex involved in transcription regulation of MHCII molecules. The RFX complex binds to the X-box DNA through the DNA binding domain of RFX5. We have examined the formation of intramolecular tyrosine cross linking, dityrosine, in RFX5DBD under oxidative stress, through UV irradiation and enzymatic action of H(2)O(2)/peroxidase by fluorescence spectroscopic studies. Dityrosine (DT) was formed predominantly in alkaline condition showing its intense characteristic fluorescence emission. Homology modeling indicated Y(39) and Y(42) could be the potential tyrosine residues undergoing oxidative cross-linking. Conformational changes in RFX5DBD under oxidative stress were observed by CD measurements. The in vitro association of X-box DNA with RFX5DBD increased DT fluorescence significantly and protected RFX5DBD from UV irradiation as observed in SDS-PAGE followed by mass spectrometric analysis. Results indicate cross protection in both RFX5DBD and DNA under oxidative stress playing important role in protein modification.
Chakraborty
Madhumita
M
Biophysics Division and Structural Genomics Section, Saha Institute of Nuclear Physics, 1/AF, Bidhannagar, Kolkata 700064, India.
Bhattacharya
Dipankar
D
Mukhopadhyay
Chaitali
C
Chakrabarti
Abhijit
A
D001881:Borates / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D018384:Oxidative Stress; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D000071701:Regulatory Factor X Transcription Factors; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000737:chemistry; D014466:Ultraviolet Rays
D001881:Borates; D004268:DNA-Binding Proteins; C093524:RFX5 protein, human; D000071701:Regulatory Factor X Transcription Factors; D014443:Tyrosine; D004247:DNA; D006861:Hydrogen Peroxide; C007543:dityrosine