Radiation damage to a DNA-binding protein. Combined circular dichroism and molecular dynamics simulation analysis

Area of Science:

• Molecular Biology
• Radiation Biology
• Biophysics

Background:

• The E. coli lactose operon is a key model for gene regulation.
• Repressor protein binding to DNA operator sites controls operon function.
• Previous studies indicated irradiation impairs repressor DNA binding.

Purpose of the Study:

• To characterize radiation-induced modifications in the lactose repressor headpiece.
• To elucidate the structural and stability changes resulting from these modifications.
• To correlate structural changes with the loss of DNA binding ability.

Main Methods:

• Mass spectrometry to identify radiation-induced lesions.
• Circular dichroism (CD) spectroscopy to assess structural changes.
• Molecular dynamics (MD) simulations of modified headpiece structures.

Main Results:

• Tyrosine residues in the headpiece are oxidized to 3,4-dihydroxyphenylalanine (DOPA) by radiation.
• CD measurements show irreversible changes in headpiece structure and stability.
• MD simulations reveal increased internal dynamics and loss of stability due to DOPA substitution.

Conclusions:

• Radiation-induced tyrosine oxidation to DOPA destabilizes the repressor headpiece.
• Altered structure and stability of the headpiece contribute to the loss of DNA binding.
• Findings are relevant to other proteins with radiosensitive amino acids in DNA-binding domains.