Vertebrate DNA crosslink repair excises toxic replication-blocking DNA crosslinks. Numerous factors involved in crosslink repair have been identified, and mutations in their corresponding genes cause Fanconi anemia (FA). A key step in crosslink repair is monoubiquitination of the FANCD2-FANCI heterodimer, which then recruits nucleases to remove the DNA lesion. In this study, monoubiquitinated FANCD2-FANCI complex was characterized using crosslinking mass spectrometry in order to provide in sights into the 3D structure of the complex.