With an affinity-purification (AP-MS) approach contact points between proteins and DNA/RNA can be uncovered. The resulting distance information is inherently of lower resolution than those provided by XL-MS, as affinity-purification deals with full peptides rather than localizable residues within peptides. The technique can however be applied to any sample complexity to discoverer DNA/RNA interacting proteins, but is most powerful to fully cover all the contact points for purified proteins/protein-complexes.
It works by ‘gluing’ the DNA/RNA to the interacting protein with formaldehyde. After proteolytic digestion, the interacting peptides remain attached to the DNA/RNA which can be enriched with IMAC. Ideally, the formaldahyde connection can be reversed and the DNA/RNA removed prior to mass spectrometry analysis. Scaling up to a full AP-MS experiment, triplicates of the sample are formaldahyde cross-linked and triplicates not treated.
From the mass spectrometry read-out a volcano plot can be constructed with the peptides enriched in the formaldahyde cross-linked samples. Knowledge of the peptides allows for annotation of existing structural models with the DNA/RNA contact points.