GBM PhD Award:
Mechanism of PINK1-mediated ubiquitin phosphorylation
Genentech, San Francisco, USA
Genetic mutations are responsible for around 10 % of all Parkinson’s disease (PD) cases. Mutations in both alleles of the PRK6 gene lead to the development of autosomal recessive juvenile parkinsonism (AR-JP). PRK6 encodes the protein kinase PINK1, which triggers the autophagy of damaged mitochondria through phosphorylation of ubiquitin (Ub) at serine 65. If this process is disrupted, for instance through loss or mutation of PINK1, damaged mitochondrial material accumulates, eventually leading to cell death.
To better understand this key event in the mitochondrial quality control pathway, we studied the PINK1-ubiquitin interaction by NMR. We could show that PINK1 targets a newly discovered confirmation of ubiquitin, in which the Ub phosphorylation site becomes exposed through a retraction of its C terminus. With this information we were able to lock Ub in this particular confirmation through specific point mutations and could generate nanobodies to stabilize the PINK1-Ub complex. Nanobody assisted crystallization yielded the first X-ray co-crystal structure of PINK1 in its active confirmation in complex with ubiquitin. This structure helped us to understand the molecular mechanism of PINK1-mediated ubiquitin phosphorylation in great detail and made it possible to explain the pathogenic effects of more than 40 mutations found in AR-JP patients.