Determining binding sites in protein-nucleic acid complexes by cross-saturation

Area of Science:

• Biophysics
• Structural Biology
• Biochemistry

Background:

• Cross-saturation experiments provide insights into interacting surfaces in protein-protein and protein-RNA complexes.
• Magnetization transfer rates are influenced by factors like geometry, spin topology, and correlation times.

Purpose of the Study:

• To assess the influence of variables on cross-saturation experiments.
• To determine the applicability range of cross-saturation for protein-nucleic acid complexes.
• To map binding surfaces without requiring deuteration.

Main Methods:

• Simulated time-course of magnetization transfer across interfaces in diverse protein-nucleic acid complexes (434 Cro, SRY, MetJ, U1A).
• Focused on experimentally accessible targets like anomeric and imino protons for selective saturation.
• Detected saturation transfer to protein moieties via reduction in HSQC spectrum peak intensities.
• Interpreted experimental cross-saturation data from Mbp1-DNA complex alongside simulations.

Main Results:

• Simulations demonstrated that cross-saturation can map protein-nucleic acid binding interfaces without deuteration.
• Information on contacting residues was obtainable by saturating nucleic acid protons.
• Experimental data from Mbp1-DNA complex corroborated simulation findings.

Conclusions:

• Cross-saturation is a valuable technique for mapping protein-nucleic acid binding interfaces.
• The method provides detailed structural information without the need for isotopic labeling or deuteration.
• This approach enhances the study of molecular interactions in biological systems.