Exchange of functional domains between a bacterial conjugative relaxase and the integrase of the human adeno-associated virus
| Autor: | Martino Bardelli, Carlos R. Escalante, Francisco Zarate-Perez, Anita F. Meier, R. Michael Linden, Matxalen Llosa, Els Henckaerts, Leticia Agúndez |
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| Přispěvatelé: | Universidad de Cantabria |
| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
BIOCHEMICAL-CHARACTERIZATION Molecular biology viruses SITE-SPECIFIC INTEGRATION lcsh:Medicine Protein domains DNA helicases Relaxase Biochemistry chemistry.chemical_compound MAMMALIAN-CELLS Materials Physics Mobile Genetic Elements GENE-PRODUCT lcsh:Science TYPE-2 REP68 PROTEIN Multidisciplinary biology Physics Genomics Dependovirus Recombinant Proteins 3. Good health Integrase Cell biology Enzymes Multidisciplinary Sciences Nucleic acids Conjugation Genetic DNA Nucleotidyltransferases Physical Sciences Science & Technology - Other Topics Helicases Sedimentation Plasmids Research Article DNA Bacterial Forms of DNA Protein domain Materials Science DNA Single-Stranded DNA construction Plasmid construction DNA replication Transfection DNA-binding protein 03 medical and health sciences Genetic Elements DNA-binding proteins Escherichia coli Genetics Humans WILD-TYPE GRAM-NEGATIVE BACTERIA Science & Technology Integrases Biology and life sciences ROLLING-CIRCLE REPLICATION lcsh:R Helicase Computational Biology Proteins DNA Endonucleases Fusion protein Research and analysis methods 030104 developmental biology HEK293 Cells Molecular biology techniques chemistry SINGLE-STRANDED-DNA PLASMID R388 biology.protein Enzymology lcsh:Q Ultracentrifugation |
| Zdroj: | PLoS One. 2018 Jul 17;13(7):e0200841 UCrea Repositorio Abierto de la Universidad de Cantabria Universidad de Cantabria (UC) Agúndez, L, Zárate-Pérez, F, Meier, A F, Bardelli, M, Llosa, M, Escalante, C R, Linden, R M & Henckaerts, E 2018, ' Exchange of functional domains between a bacterial conjugative relaxase and the integrase of the human adeno-associated virus ', PLoS ONE, vol. 13, no. 7, e0200841 . https://doi.org/10.1371/journal.pone.0200841 PLoS ONE PLoS ONE, Vol 13, Iss 7, p e0200841 (2018) Digital.CSIC. Repositorio Institucional del CSIC instname |
| Popis: | © 2018 Agúndez et al. Endonucleases of the HUH family are specialized in processing single-stranded DNA in a variety of evolutionarily highly conserved biological processes related to mobile genetic elements. They share a structurally defined catalytic domain for site-specific nicking and strand-transfer reactions, which is often linked to the activities of additional functional domains, contributing to their overall versatility. To assess if these HUH domains could be interchanged, we created a chimeric protein from two distantly related HUH endonucleases, containing the N-terminal HUH domain of the bacterial conjugative relaxase TrwC and the C-terminal DNA helicase domain of the human adeno-associated virus (AAV) replicase and site-specific integrase. The purified chimeric protein retained oligomerization properties and DNA helicase activities similar to Rep68, while its DNA binding specificity and cleaving-joining activity at oriT was similar to TrwC. Interestingly, the chimeric protein could catalyse site-specific integration in bacteria with an efficiency comparable to that of TrwC, while the HUH domain of TrwC alone was unable to catalyze this reaction, implying that the Rep68 C-terminal helicase domain is complementing the TrwC HUH domain to achieve site-specific integration into TrwC targets in bacteria. Our results illustrate how HUH domains could have acquired through evolution other domains in order to attain new roles, contributing to the functional flexibility observed in this protein superfamily. This work was supported by the Medical Research Council (MRC) grant MR/N022890/1 to EH and grant 1001764 to RML; National Institutes of Health (NIH) grant RO1-GM09285 to CRE; Spanish Ministry of Economy and Competitiveness (MINECO) grant BIO2013-46414-P to ML and AFM is supported by a Doc.Mobility fellowship from the Swiss National Science Foundation. |
| Databáze: | OpenAIRE |
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