BRCA1 RING domain cancer-predisposing mutations - Structural consequences and effects on protein-protein interactions

Cancer-predisposing missense mutations in the RING domain of BRCA1 primaril y target Zn2+-liganding residues. Here we report on the structural conseque nces of such mutations introduced into the second Zn2+ site (Site II) of th e BRCA1 RING domain and their effect on the interaction with the BARD1 RING domain. Each of the BRCA1 Site II mutants still interact and form a stable heterodimer with BARD1. Limited proteolysis of BRCA1/BARD1 complexes, moni tored by matrix-assisted laser desorption ionization time-of-flight spectro metry, show that the mutations cause a local structural perturbation that i s primarily confined to the second Zn2+ binding loop of the BRCA1 subunit. These findings are consistent with the structure of the BRCA1/BARD1 heterod imer, which shows this region is well removed from the helices required for dimerization with BARD1. Instead, the mutations alter a region of BRCA1 th at appears to be required for interaction with ubiquitin-conjugating enzyme s.