{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,4]],"date-time":"2026-06-04T02:08:06Z","timestamp":1780538886407,"version":"3.54.1"},"reference-count":30,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2026,6,1]],"date-time":"2026-06-01T00:00:00Z","timestamp":1780272000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biomolecules"],"abstract":"<jats:p>Babesia microti is the primary agent of human babesiosis, an emerging tick-borne disease with limited treatment options and growing evidence of drug resistance. Deubiquitinases (DUBs) play critical roles in protein homeostasis and host\u2013pathogen interactions, yet none have been characterized in B. microti. Here, we report the first molecular cloning, expression, and functional characterization of an OTU-like cysteine protease from B. microti (bm-OTU). The recombinant bm-OTU protein (~22 kDa) was expressed in E. coli, purified to high homogeneity, and exhibited ~93% solubility under native conditions. In vitro fluorogenic assays confirmed its deubiquitinase activity. Expression of bm-OTU in HEK293T cells was associated with reduced ubiquitination in cells and increased apoptosis in this overexpression model, as demonstrated by flow cytometry and Western blot analyses. Furthermore, transcriptomic analysis revealed that bm-OTU modulates host immune pathways, notably suppressing the expression of interferon-stimulated genes (APOBEC3G, G1P2) while upregulating the pro-apoptotic gene BAK and the inflammasome sensor AIM2. These findings establish bm-OTU as a functional deubiquitinase and a potential virulence factor that may contribute to immune evasion and pathogenesis in babesiosis and may represent a potential target, pending further validation.<\/jats:p>","DOI":"10.3390\/biom16060819","type":"journal-article","created":{"date-parts":[[2026,6,4]],"date-time":"2026-06-04T01:08:02Z","timestamp":1780535282000},"page":"819","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Characterization of a Novel OTU-like Deubiquitinase from Babesia microti: Implications for Babesiosis Treatment"],"prefix":"10.3390","volume":"16","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2721-4004","authenticated-orcid":false,"given":"Bet\u00fcl","family":"Yusuf","sequence":"first","affiliation":[{"name":"College of Engineering, American University of Sharjah, University City, Sharjah 26666, United Arab Emirates"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8096-6056","authenticated-orcid":false,"given":"Fatih","family":"Kocaba\u015f","sequence":"additional","affiliation":[{"name":"Department of Molecular Biology and Genetics, Faculty of Engineering and Natural Sciences, Istanbul Atlas University, Istanbul 34403, T\u00fcrkiye"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,1]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Vannier, E., and Krause, P.J. (2020). Babesiosis. Hunter\u2019s Tropical Medicine and Emerging Infectious Diseases, Elsevier.","DOI":"10.1016\/B978-0-323-55512-8.00105-8"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Yang, Y., Christie, J., K\u00f6ster, L., Du, A., and Yao, C. (2021). Emerging Human Babesiosis with \u201cGround Zero\u201d in North America. Microorganisms, 9.","DOI":"10.3390\/microorganisms9020440"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"14","DOI":"10.1128\/CMR.00022-10","article-title":"Transfusion-transmitted Babesia spp.: Bull\u2019s-eye on Babesia microti","volume":"24","author":"Leiby","year":"2011","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"165","DOI":"10.1016\/j.ijpara.2018.11.007","article-title":"Human babesiosis","volume":"49","author":"Krause","year":"2019","journal-title":"Int. J. Parasitol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"2149","DOI":"10.1001\/archinte.158.19.2149","article-title":"Human babesiosis in New York State: Review of 139 hospitalized cases and analysis of prognostic factors","volume":"158","author":"White","year":"1998","journal-title":"Arch. Intern. Med."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1454","DOI":"10.1056\/NEJM200011163432004","article-title":"Atovaquone and azithromycin for the treatment of babesiosis","volume":"343","author":"Krause","year":"2000","journal-title":"N. Engl. J. Med."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"370","DOI":"10.1086\/525852","article-title":"Persistent and relapsing babesiosis in immunocompromised patients","volume":"46","author":"Krause","year":"2008","journal-title":"Clin. Infect. Dis."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Florin-Christensen, M., Wieser, S.N., Suarez, C.E., and Schnittger, L. (2021). In silico survey and characterization of Babesia microti functional and non-functional proteases. Pathogens, 10.","DOI":"10.3390\/pathogens10111457"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"2727","DOI":"10.1073\/pnas.95.6.2727","article-title":"The ubiquitin-proteasome pathway: The complexity and myriad functions of proteins death","volume":"95","author":"Ciechanover","year":"1998","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_10","first-page":"113","article-title":"Ubiquitin recognition by the proteasome","volume":"161","author":"Saeki","year":"2017","journal-title":"J. Biochem."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"550","DOI":"10.1038\/nrm2731","article-title":"Breaking the chains: Structure and function of the deubiquitinases","volume":"10","author":"Komander","year":"2009","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"3815","DOI":"10.1128\/JVI.03252-12","article-title":"Diversity of ubiquitin and ISG15 specificity among nairoviruses\u2019 viral ovarian tumor domain proteases","volume":"87","author":"Capodagli","year":"2013","journal-title":"J. Virol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"2228","DOI":"10.1073\/pnas.1015287108","article-title":"Molecular basis for ubiquitin and ISG15 cross-reactivity in viral ovarian tumor domains","volume":"108","author":"Akutsu","year":"2011","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"190","DOI":"10.1007\/s11262-015-1226-5","article-title":"Fluorometric CCHFV OTU protease assay with potent inhibitors","volume":"51","author":"Kocabas","year":"2015","journal-title":"Virus Genes"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"239","DOI":"10.3906\/biy-1501-56","article-title":"Identification of small molecule binding pocket for inhibition of Crimean-Congo hemorrhagic fever virus OTU protease","volume":"40","author":"Kocabas","year":"2016","journal-title":"Turk. J. Biol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"4155","DOI":"10.1002\/jps.21327","article-title":"Measuring and increasing protein solubility","volume":"97","author":"Trevino","year":"2008","journal-title":"J. Pharm. Sci."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"501","DOI":"10.1038\/nri2337","article-title":"Deubiquitylation and regulation of the immune response","volume":"8","author":"Sun","year":"2008","journal-title":"Nat. Rev. Immunol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"439","DOI":"10.1038\/nrm3143","article-title":"Ubiquitylation in apoptosis: DUBs to the rescue","volume":"12","author":"Vucic","year":"2011","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"567","DOI":"10.1146\/annurev-virology-092818-015756","article-title":"Interferon-stimulated genes: What do they all do?","volume":"6","author":"Schoggins","year":"2019","journal-title":"Annu. Rev. Virol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"404","DOI":"10.1016\/j.chom.2007.09.014","article-title":"Ovarian tumor domain-containing viral proteases evade ubiquitin- and ISG15-dependent innate immune responses","volume":"2","author":"Giannakopoulos","year":"2007","journal-title":"Cell Host Microbe"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"509","DOI":"10.1128\/CMR.17.3.509-539.2004","article-title":"Pathogenesis of malaria and clinically similar conditions","volume":"17","author":"Clark","year":"2004","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_22","first-page":"81","article-title":"Seroprevalance of Babesia microti in humans living in rural areas of the Sinop region","volume":"34","author":"Poyraz","year":"2010","journal-title":"Turk. Parazitolojii Derg."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Ozubek, S., Bastos, R.G., Alzan, H.F., Inci, A., Aktas, M., and Suarez, C.E. (2020). Bovine babesiosis in Turkey: Impact, current gaps, and opportunities for intervention. Pathogens, 9.","DOI":"10.3390\/pathogens9121041"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"3445","DOI":"10.1042\/BCJ20210481","article-title":"Identification of first-in-class plasmodium OTU inhibitors with potent anti-malarial activity","volume":"478","author":"Siyah","year":"2021","journal-title":"Biochem. J."},{"key":"ref_25","first-page":"100100","article-title":"Immune evasion strategies of Seneca Valley virus: Mechanisms of host innate immune suppression","volume":"3","author":"Xie","year":"2025","journal-title":"Agric. Commun."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"875","DOI":"10.1111\/zph.12983","article-title":"Zoonotic Babesia microti infection in wild rodents in Erzurum province, northeastern Turkey","volume":"69","author":"Guven","year":"2022","journal-title":"Zoonoses Public Health"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"65","DOI":"10.1007\/s00436-014-4160-x","article-title":"Molecular identification of Theileria and Babesia in ticks collected from sheep and goats in the Black Sea region of Turkey","volume":"114","author":"Aydin","year":"2015","journal-title":"Parasitol. Res."},{"key":"ref_28","first-page":"39","article-title":"Babesiosis as a potential hazard in animal production and its global emerging zoonotic threat for One Health","volume":"2","author":"Ijaz","year":"2024","journal-title":"Agro Sci. J. Igdir Univ."},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Wehrmann, M., and Vilchez, D. (2023). The emerging role and therapeutic implications of bacterial and parasitic deubiquitinating enzymes. Front. Immunol., 14.","DOI":"10.3389\/fimmu.2023.1303072"},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Kumar, P., Kumar, P., Mandal, D., and Velayutham, R. (2022). The emerging role of Deubiquitinases (DUBs) in parasites: A foresight review. Front. Cell. Infect. Microbiol., 12.","DOI":"10.3389\/fcimb.2022.985178"}],"container-title":["Biomolecules"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2218-273X\/16\/6\/819\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,4]],"date-time":"2026-06-04T01:49:23Z","timestamp":1780537763000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2218-273X\/16\/6\/819"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,6,1]]},"references-count":30,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2026,6]]}},"alternative-id":["biom16060819"],"URL":"https:\/\/doi.org\/10.3390\/biom16060819","relation":{},"ISSN":["2218-273X"],"issn-type":[{"value":"2218-273X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,6,1]]}}}