Role of conformational dynamics in sequence-specific autoantibody*ssDNA recognition

Area of Science:

• Immunology
• Structural Biology
• Molecular Biophysics

Background:

• Monoclonal autoantibody 11F8 targets single-stranded DNA (ssDNA).
• It originates from lupus-prone mice and contributes to kidney damage.
• Specificity is linked to a somatic mutation (serine to arginine at 31V(H)).

Purpose of the Study:

• Investigate the role of the 31V(H) mutation in 11F8 antibody affinity and specificity.
• Elucidate the structural and thermodynamic basis of ssDNA recognition.
• Characterize conformational changes upon antibody-ssDNA complex formation.

Main Methods:

• Site-directed mutagenesis to revert the 31V(H) mutation.
• Affinity, thermodynamic, and kinetic measurements.
• Fluorescence resonance energy transfer (FRET) to study conformational dynamics.

Main Results:

• Reversion to germline serine decreased 11F8 affinity for ssDNA by over 30-fold.
• The mutated arginine (R31V(H)) forms key interactions (salt bridge, hydrogen bond) with ssDNA.
• Greater conformational rearrangement in five of six complementarity-determining regions was observed with the mutated arginine.

Conclusions:

• The somatic mutation to arginine at 31V(H) is critical for high-affinity ssDNA binding and pathogenic potential.
• This mutation induces significant conformational changes in the antibody binding site.
• Understanding these structural dynamics provides insights into lupus autoimmunity and antibody engineering.