Extended Data Fig. 7: The phospho-Ubl binding site on the UPD.

a, Side-by-side view of phospho-parkin–phospho-ubiquitin (left) and parkin–phospho-ubiquitin (PDB 5N2W⁶, right), and superposition of both (below). The green Ubl domain changes position by >50 Å. b, E2–Ub from the structure of the HOIP RBR domain in complex with UBE2D2–Ub²⁸ was modelled onto phospho-parkin–phospho-ubiquitin, by superposition of the RING1 domains of each complex. The E2-conjugated ubiquitin molecule in the ‘open’ conformation binds to the previously recognized cryptic ubiquitin binding interface on RING1–IBR⁶. The contact points correlate with HDX-MS data (Fig. 1d, Extended Data Figs. 2, 3c). c, HDX-MS data from Fig. 1e were plotted onto the phospho-parkin–phospho-ubiquitin structure with identical colouring (blue, more protected from solvent exchange; red, less protected from solvent exchange; grey, not covered in all of the compared states, compare with Fig. 1). Protected regions on UPD match the observed phospho-Ubl interface. d, HDX-MS experiments comparing parkin with a mutation in the phospho-acceptor binding site on the UPD (phospho-parkin(K211N)–phospho-ubiquitin) compared with phospho-parkin–phospho-ubiquitin, coloured as in c. The mutant is unable to protect the Ubl, and to release RING2 and REP. Experiments were done as technical triplicate.