Mechanism of feedforward Parkin activation by self-association

Abstract Loss of function Parkin mutations lead to early-onset of Parkinson’s disease. Parkin is an auto-inhibited ubiquitin E3-ligase activated by phosphorylation of its ubiquitin-like (Ubl) domain and ubiquitin by PINK1. Herein, we show a competitive binding mode of the phospho-Ubl and RING2 domains on the RING0 domain, which regulates Parkin activity. We show that phosphorylated Parkin can complex with unmodified Parkin, leading to the activation of autoinhibited Parkin in trans. Furthermore, we show that the activator element (ACT) of Parkin is required to maintain the enzyme’s kinetics, and the removal of ACT slows the enzyme catalysis. We also demonstrate that ACT can activate Parkin in trans but less efficiently than when present in the cis molecule. Furthermore, we have identified and characterized a new donor ubiquitin binding pocket that plays a crucial role in function. This study provides new molecular insights into Parkin activation which may aid the development of Parkin activators as a therapeutic strategy against Parkinson’s.