Affiliation:
1. Walter and Eliza Hall Institute of Medical Research
2. Bio21 Molecular Science and Biotechnology Institute, Parkville, Australia; and Department of Biochemistry and Pharmacology, The University of Melbourne, Melbourne, Australia
Abstract
The phylum Apicomplexa contains several parasitic species of medical and agricultural importance. The ubiquitination machinery remains, for the most part, uncharacterised in apicomplexan parasites, despite the important roles that it plays in eukaryotic biology. Bioinformatic analysis of the ubiquitination machinery in apicomplexan parasites revealed an expanded ovarian tumour domain–containing (OTU) deubiquitinase (DUB) family inToxoplasma, potentially reflecting functional importance in apicomplexan parasites. This study presents comprehensive characterisation ofToxoplasmaOTU DUBs. AlphaFold-guided structural analysis not only confirmed functional orthologues found across eukaryotes, but also identified apicomplexan-specific enzymes, subsequently enabling discovery of a cryptic OTU DUB inPlasmodiumspecies. Comprehensive biochemical characterisation of 11ToxoplasmaOTU DUBs revealed activity against ubiquitin- and NEDD8-based substrates and revealed ubiquitin linkage preferences for Lys6-, Lys11-, Lys48-, and Lys63-linked chain types. We show that accessory domains inToxoplasmaOTU DUBs impose linkage preferences, and in case of apicomplexan-specific TgOTU9, we discover a cryptic ubiquitin-binding domain that is essential for TgOTU9 activity. Using the auxin-inducible degron (AID) to generate knockdown parasite lines, TgOTUD6B was found to be important forToxoplasmagrowth.
Funder
DHAC | National Health and Medical Research Council
Walter and Eliza Hall Institute
Australian Government
Victorian State Government
Publisher
Life Science Alliance, LLC
Subject
Health, Toxicology and Mutagenesis,Plant Science,Biochemistry, Genetics and Molecular Biology (miscellaneous),Ecology
Cited by
2 articles.
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