Structural ubiquitin contributes to K48 linkage specificity of the HECT ligase Tom1
Katrina Warner,
Moritz Hunkeler,
Kheewoong Baek,
Anna Schmoker,
Shourya S. Roy Burman,
Daan Overwijn,
Cyrus Jin,
Katherine A. Donovan,
Eric S. Fischer
Affiliations
Katrina Warner
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02215, USA
Moritz Hunkeler
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02215, USA
Kheewoong Baek
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02215, USA
Anna Schmoker
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02215, USA
Shourya S. Roy Burman
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02215, USA
Daan Overwijn
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA
Cyrus Jin
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02215, USA
Katherine A. Donovan
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02215, USA
Eric S. Fischer
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02215, USA; Corresponding author
Summary: Homologous to E6AP C terminus (HECT) ubiquitin ligases play key roles in essential pathways such as DNA repair, cell cycle control, or protein quality control. Tom1 is one of five HECT ubiquitin E3 ligases in budding yeast S. cerevisiae and is prototypical for a ligase with pleiotropic functions such as ubiquitin chain amplification, orphan quality control, and DNA damage response. Structures of full-length HECT ligases, including the Tom1 ortholog HUWE1, have been reported, but how domains beyond the conserved catalytic module contribute to catalysis remains largely elusive. Here, through cryoelectron microscopy (cryo-EM) snapshots of Tom1 during an active ubiquitination cycle, we demonstrate that the extended domain architecture directly contributes to activity. We identify a Tom1-ubiquitin architecture during ubiquitination involving a non-canonical ubiquitin-binding site in the solenoid shape of Tom1. We demonstrate that this ubiquitin-binding site coordinates a structural ubiquitin contributing to the fidelity of K48 poly-ubiquitin chain assembly.
