Proximity-enhanced SuFEx chemical cross-linker for specific and multitargeting cross-linking mass spectrometry

Significance Chemical cross-linking mass spectrometry is a powerful method to identify protein interaction partners. The cross-links also provide approximate interresidue distances, which can help model the structures of complexes. However, current cross-linking reagents react with only a limited set of amino acid residues and their very high reactivity can induce cross-links between sites that are distant in the native state of the interrogated proteins. We therefore developed a “plant-and-cast” strategy, in which a heterobifunctional cross-linker is “planted” onto surface Lys residues using a highly reactive succinimide ester. The half-reacted cross-linker then “casts” a much less reactive sulfonyl fluoride across the proximal surface resulting in cross-links to neighboring Ser, Thr, Tyr, His, or Lys sidechains in a proximity-enhanced reaction.