characterisation of the trichinella spiralis deubiquitinating enzyme, tsuch37, an evolutionarily conserved proteasome interaction partner描述的旋毛虫deubiquitinating酶、tsuch37一个进化守恒的蛋白酶体的交互合作伙伴.pdfVIP

Characterisation of the Trichinella spiralis Deubiquitinating Enzyme, TsUCH37, an Evolutionarily Conserved Proteasome Interaction Partner 1 1 2 3 1 Rhiannon R. White , Sachiko Miyata , Eliseo Papa , Eric Spooner , Kleoniki Gounaris , Murray E. 1 1 Selkirk , Katerina Artavanis-Tsakonas * 1 Division of Cell and Molecular Biology, Department of Life Sciences, Imperial College London, London, United Kingdom, 2 Health Sciences and Technology, Harvard/ Massachusetts Institute of Technology (MIT), Cambridge, Massachusetts, United States of America, 3 Whitehead Institute for Biomedical Research, Cambridge, Massachusetts, United States of America Abstract Background: Trichinella spiralis is a zoonotic parasitic nematode that causes trichinellosis, a disease that has been identified on all continents except Antarctica. During chronic infection, T. spiralis larvae infect skeletal myofibres, severely disrupting their differentiation state. Methodology and Results: An activity-based probe, HA-Ub-VME, was used to identify deubiquitinating enzyme (DUB) activity in lysate of T. spiralis L1 larvae. Results were analysed by immuno-blot and immuno-precipitation, identifying a number of potential DUBs. Immuno-precipitated proteins were subjected to LC/MS/MS, yielding peptides with sequence homology to 5 conserved human DUBs: UCH-L5, UCH-L3, HAUSP, OTU 6B and Ataxin-3. The predicted gene encoding the putative UCH-L5 homologue, TsUCH37, was cloned and recombinant protein was expressed and purified. The deubiquitinating activity of this enzyme was verified by Ub-AMC assay. Co-precipitation of recombinant TsUCH37 showed that the protein associate