Structure of PINK1 in complex with its substrate ubiquitin

Autosomal-recessive juvenile Parkinsonism (AR-JP) is caused by mutations in a number of PARK genes, in particular the genes encoding the E3 ubiquitin ligase Parkin (PARK2, also known as PRKN) and its upstream protein kinase PINK1 (also known as PARK6). PINK1 phosphorylates both ubiquitin and the ubiquitin-like domain of Parkin on structurally protected Ser65 residues, triggering mitophagy. Here we report a crystal structure of a nanobody-stabilized complex containing Pediculus humanus corporis (Ph)PINK1 bound to ubiquitin in the ‘C-terminally retracted’ (Ub-CR) conformation. The structure reveals many peculiarities of PINK1, including the architecture of the C-terminal region, and reveals how the N lobe of PINK1 binds ubiquitin via a unique insertion. The flexible Ser65 loop in the Ub-CR conformation contacts the activation segment, facilitating placement of Ser65 in a phosphate-accepting position. The structure also explains how autophosphorylation in the N lobe stabilizes structurally and functionally important insertions, and reveals the molecular basis of AR-JP-causing mutations, some of which disrupt ubiquitin binding. Stabilization of a transient protein kinase–substrate complex using a nanobody provides molecular details about how the Parkinson’s disease-linked protein kinase PINK1 phosphorylates ubiquitin, and suggests new pharmacological strategies. The kinase enzyme PINK1 is known mainly for two reasons. At an organism level, mutations of PINK1 have been associated with autosomal-recessive juvenile Parkinsonism (AR-JP). At a cellular level, PINK1 phosphorylates both ubiquitin and a ubiquitin-like domain within its partner enzyme Parkin to trigger mitophagy, the process by which cells get rid of dysfunctional mitochondria. David Komander and co-authors report the structure of a complex between louse PINK1 and ubiquitin, which they obtained using nanobody-based stabilization. The structure provides molecular insights not only into PINK1–ubiquitin interactions and therefore the mechanism of PINK1 activity, but also into AR-JP-associated mutations, some of which disrupt ubiquitin binding.