Intricate mechanism (s) of substrate specificity and loss of function on disease mutation (K211N) of Parkin

PINK1 and Parkin mutations lead to the early onset of Parkinsons disease. PINK1-mediated phosphorylation of its substrates such as ubiquitin (Ub), ubiquitin-like protein (NEDD8), and ubiquitin-like (Ubl) domain of Parkin activate autoinhibited Parkin E3 ligase. The mechanism of various phospho-Ubls binding on Parkin and conformational changes leading to Parkin activation remain elusive. Herein, we determine the first crystal structure of human Parkin E3 ligase bound with phospho (p)-NEDD8, which shows that NEDD8 has evolved over Ub to bind and activate Parkin more robustly. X-ray crystal structures and supporting biophysical/biochemical data reveal specific recognition and underlying mechanisms of pUb/pNEDD8 and pUbl domain binding to the RING1 and RING0 domains, respectively. This new data also shows that pUb/pNEDD8 binding in the RING1 pocket causes allosteric conformational changes in Parkins catalytic domain (RING2), leading to Parkin activation. Furthermore, Parkinsons disease mutation K211N in the RING0 domain of Parkin was believed to lose its activation due to loss of interaction with pUb. However, our data reveal allosteric conformational changes due to N211 that lock RING2 with RING0 to inhibit Parkin K211N activity without disrupting pNEDD8/pUb binding. This study would aid the design of small-molecule Parkin activators for the treatment of Parkinsons disease.