2 research outputs found
Synergistic recruitment of UbcH7~Ub and phosphorylated Ubl domain triggers parkin activation
The E3 ligase parkin ubiquitinates outer mitochondrial membrane
proteins during oxidative stress and is linked to early-onset
Parkinson’s disease. Parkin is autoinhibited but is activated by the
kinase PINK1 that phosphorylates ubiquitin leading to parkin
recruitment, and stimulates phosphorylation of parkin’s N-terminal
ubiquitin-like (pUbl) domain. How these events alter the
structure of parkin to allow recruitment of an E2~Ub conjugate
and enhanced ubiquitination is an unresolved question. We
present a model of an E2~Ub conjugate bound to the phosphoubiquitin-loaded
C-terminus of parkin, derived from NMR chemical
shift perturbation experiments. We show the UbcH7~Ub conjugate
binds in the open state whereby conjugated ubiquitin binds to the
RING1/IBR interface. Further, NMR and mass spectrometry experiments
indicate the RING0/RING2 interface is re-modelled,
remote from the E2 binding site, and this alters the reactivity of
the RING2(Rcat) catalytic cysteine, needed for ubiquitin transfer.
Our experiments provide evidence that parkin phosphorylation
and E2~Ub recruitment act synergistically to enhance a weak
interaction of the pUbl domain with the RING0 domain and rearrange
the location of the RING2(Rcat) domain to drive parkin
activity
Structural Basis for the Inhibition of Host Protein Ubiquitination by Shigella Effector Kinase OspG
SummaryShigella invasion of its human host is assisted by T3SS-delivered effector proteins. The OspG effector kinase binds ubiquitin and ubiquitin-loaded E2-conjugating enzymes, including UbcH5b and UbcH7, and attenuates the host innate immune NF-kB signaling. We present the structure of OspG bound to the UbcH7∼Ub conjugate. OspG has a minimal kinase fold lacking the activation loop of regulatory kinases. UbcH7∼Ub binds OspG at sites remote from the kinase active site, yet increases its kinase activity. The ubiquitin is positioned in the “open” conformation with respect to UbcH7 using its I44 patch to interact with the C terminus of OspG. UbcH7 binds to OspG using two conserved loops essential for E3 ligase recruitment. The interaction of the UbcH7∼Ub with OspG is remarkably similar to the interaction of an E2∼Ub with a HECT E3 ligase. OspG interferes with the interaction of UbcH7 with the E3 parkin and inhibits the activity of the E3