Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation
| Autor(a) principal: | |
|---|---|
| Data de Publicação: | 2023 |
| Outros Autores: | , , , , , , , , |
| Tipo de documento: | Artigo |
| Idioma: | eng |
| Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
| Texto Completo: | http://hdl.handle.net/10362/158545 |
Resumo: | Funding Information: This work was supported by grants of Universidad de la República (CSIC_2018 and EI_2020) to R. R., Universidad de la República (CSIC Iniciación_2017_ID_159) to N. C., Alexander von Humboldt Foundation (AvHF) to T. G. and R. R., the Novo Nordisk Foundation ( NNF13OC0004294 and NNF20SA0064214 ) to M. J. D., and PICT 2018-0795 from Agencia I+d+i Argentina , University of Buenos Aires (grant 20020120300025BA ), and CONICET (grant 11220150100303CO ) to D. E. N. C. was partially supported by a fellowship from Comisión Académica de Posgrado (CAP), Universidad de la República , Uruguay. Additional funding was obtained from Programa de Desarrollo de las Ciencias Básicas (PEDECIBA, Uruguay), Agencia Nacional de Investigación e Innovación (ANII-SNI, Uruguay), EU-LAC Health (EULACH16/T01-0131), and Programa de Alimentos y Salud Humana (PAyS, Uruguay). Funding Information: The authors would like to thank Dr Tobias Karlberg and Dr Susanne Gräslund from the Karolinska Institutet - Structural Genomics Consortium for providing the HsGS plasmid (Construct ID GLULA-c004). We also thank Dr Verónica Tórtora for her major aid within the initial steps of the transformation and HsGS expression experiments. N. C. D. E. P. H. T. G. M. J. D. S. B. and R. R. conceptualization; N. C. Mauricio Mastrogiovanni, Michele Mariotti, F. M. I. and P. H. methodology; N. C. Mauricio Mastrogiovanni, Michele Mariotti, and F. M. I. investigation; N. C. writing–original draft; M. J. D. S. B. and R. R. writing–review and editing; R. R. supervision. This work was supported by grants of Universidad de la República (CSIC_2018 and EI_2020) to R. R. Universidad de la República (CSIC Iniciación_2017_ID_159) to N. C. Alexander von Humboldt Foundation (AvHF) to T. G. and R. R. the Novo Nordisk Foundation (NNF13OC0004294 and NNF20SA0064214) to M. J. D. and PICT 2018-0795 from Agencia I+d+i Argentina, University of Buenos Aires (grant 20020120300025BA), and CONICET (grant 11220150100303CO) to D. E. N. C. was partially supported by a fellowship from Comisión Académica de Posgrado (CAP), Universidad de la República, Uruguay. Additional funding was obtained from Programa de Desarrollo de las Ciencias Básicas (PEDECIBA, Uruguay), Agencia Nacional de Investigación e Innovación (ANII-SNI, Uruguay), EU-LAC Health (EULACH16/T01-0131), and Programa de Alimentos y Salud Humana (PAyS, Uruguay). Publisher Copyright: © 2023 The Authors |
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Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregationaggregationdityrosinefree radicalsglutamine synthetasehydrogen peroxidenitrotyrosineoxidantsperoxynitritethiol oxidationBiochemistryMolecular BiologyCell BiologySDG 3 - Good Health and Well-beingFunding Information: This work was supported by grants of Universidad de la República (CSIC_2018 and EI_2020) to R. R., Universidad de la República (CSIC Iniciación_2017_ID_159) to N. C., Alexander von Humboldt Foundation (AvHF) to T. G. and R. R., the Novo Nordisk Foundation ( NNF13OC0004294 and NNF20SA0064214 ) to M. J. D., and PICT 2018-0795 from Agencia I+d+i Argentina , University of Buenos Aires (grant 20020120300025BA ), and CONICET (grant 11220150100303CO ) to D. E. N. C. was partially supported by a fellowship from Comisión Académica de Posgrado (CAP), Universidad de la República , Uruguay. Additional funding was obtained from Programa de Desarrollo de las Ciencias Básicas (PEDECIBA, Uruguay), Agencia Nacional de Investigación e Innovación (ANII-SNI, Uruguay), EU-LAC Health (EULACH16/T01-0131), and Programa de Alimentos y Salud Humana (PAyS, Uruguay). Funding Information: The authors would like to thank Dr Tobias Karlberg and Dr Susanne Gräslund from the Karolinska Institutet - Structural Genomics Consortium for providing the HsGS plasmid (Construct ID GLULA-c004). We also thank Dr Verónica Tórtora for her major aid within the initial steps of the transformation and HsGS expression experiments. N. C. D. E. P. H. T. G. M. J. D. S. B. and R. R. conceptualization; N. C. Mauricio Mastrogiovanni, Michele Mariotti, F. M. I. and P. H. methodology; N. C. Mauricio Mastrogiovanni, Michele Mariotti, and F. M. I. investigation; N. C. writing–original draft; M. J. D. S. B. and R. R. writing–review and editing; R. R. supervision. This work was supported by grants of Universidad de la República (CSIC_2018 and EI_2020) to R. R. Universidad de la República (CSIC Iniciación_2017_ID_159) to N. C. Alexander von Humboldt Foundation (AvHF) to T. G. and R. R. the Novo Nordisk Foundation (NNF13OC0004294 and NNF20SA0064214) to M. J. D. and PICT 2018-0795 from Agencia I+d+i Argentina, University of Buenos Aires (grant 20020120300025BA), and CONICET (grant 11220150100303CO) to D. E. N. C. was partially supported by a fellowship from Comisión Académica de Posgrado (CAP), Universidad de la República, Uruguay. Additional funding was obtained from Programa de Desarrollo de las Ciencias Básicas (PEDECIBA, Uruguay), Agencia Nacional de Investigación e Innovación (ANII-SNI, Uruguay), EU-LAC Health (EULACH16/T01-0131), and Programa de Alimentos y Salud Humana (PAyS, Uruguay). Publisher Copyright: © 2023 The AuthorsGlutamine synthetase (GS), which catalyzes the ATP-dependent synthesis of L-glutamine from L-glutamate and ammonia, is a ubiquitous and conserved enzyme that plays a pivotal role in nitrogen metabolism across all life domains. In vertebrates, GS is highly expressed in astrocytes, where its activity sustains the glutamate-glutamine cycle at glutamatergic synapses and is thus essential for maintaining brain homeostasis. In fact, decreased GS levels or activity have been associated with neurodegenerative diseases, with these alterations attributed to oxidative post-translational modifications of the protein, in particular tyrosine nitration. In this study, we expressed and purified human GS (HsGS) and performed an in-depth analysis of its oxidative inactivation by peroxynitrite (ONOO−) in vitro. We found that ONOO− exposure led to a dose-dependent loss of HsGS activity, the oxidation of cysteine, methionine, and tyrosine residues and also the nitration of tryptophan and tyrosine residues. Peptide mapping by LC-MS/MS through combined H216O/H218O trypsin digestion identified up to 10 tyrosine nitration sites and five types of dityrosine cross-links; these modifications were further scrutinized by structural analysis. Tyrosine residues 171, 185, 269, 283, and 336 were the main nitration targets; however, tyrosine-to-phenylalanine HsGS mutants revealed that their sole nitration was not responsible for enzyme inactivation. In addition, we observed that ONOO− induced HsGS aggregation and activity loss. Thiol oxidation was a key modification to elicit aggregation, as it was also induced by hydrogen peroxide treatment. Taken together, our results indicate that multiple oxidative events at various sites are responsible for the inactivation and aggregation of human GS.Instituto de Tecnologia Química e Biológica António Xavier (ITQB)RUNCampolo, NicolásMastrogiovanni, MauricioMariotti, MicheleIssoglio, Federico M.Estrin, DaríoHägglund, PerGrune, TilmanDavies, Michael J.Bartesaghi, SilvinaRadi, Rafael2023-09-30T22:21:36Z2023-032023-03-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10362/158545eng0021-9258PURE: 72625831https://doi.org/10.1016/j.jbc.2023.102941info:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2024-03-11T05:41:05Zoai:run.unl.pt:10362/158545Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T03:57:12.338265Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse |
| dc.title.none.fl_str_mv |
Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation |
| title |
Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation |
| spellingShingle |
Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation Campolo, Nicolás aggregation dityrosine free radicals glutamine synthetase hydrogen peroxide nitrotyrosine oxidants peroxynitrite thiol oxidation Biochemistry Molecular Biology Cell Biology SDG 3 - Good Health and Well-being |
| title_short |
Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation |
| title_full |
Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation |
| title_fullStr |
Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation |
| title_full_unstemmed |
Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation |
| title_sort |
Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation |
| author |
Campolo, Nicolás |
| author_facet |
Campolo, Nicolás Mastrogiovanni, Mauricio Mariotti, Michele Issoglio, Federico M. Estrin, Darío Hägglund, Per Grune, Tilman Davies, Michael J. Bartesaghi, Silvina Radi, Rafael |
| author_role |
author |
| author2 |
Mastrogiovanni, Mauricio Mariotti, Michele Issoglio, Federico M. Estrin, Darío Hägglund, Per Grune, Tilman Davies, Michael J. Bartesaghi, Silvina Radi, Rafael |
| author2_role |
author author author author author author author author author |
| dc.contributor.none.fl_str_mv |
Instituto de Tecnologia Química e Biológica António Xavier (ITQB) RUN |
| dc.contributor.author.fl_str_mv |
Campolo, Nicolás Mastrogiovanni, Mauricio Mariotti, Michele Issoglio, Federico M. Estrin, Darío Hägglund, Per Grune, Tilman Davies, Michael J. Bartesaghi, Silvina Radi, Rafael |
| dc.subject.por.fl_str_mv |
aggregation dityrosine free radicals glutamine synthetase hydrogen peroxide nitrotyrosine oxidants peroxynitrite thiol oxidation Biochemistry Molecular Biology Cell Biology SDG 3 - Good Health and Well-being |
| topic |
aggregation dityrosine free radicals glutamine synthetase hydrogen peroxide nitrotyrosine oxidants peroxynitrite thiol oxidation Biochemistry Molecular Biology Cell Biology SDG 3 - Good Health and Well-being |
| description |
Funding Information: This work was supported by grants of Universidad de la República (CSIC_2018 and EI_2020) to R. R., Universidad de la República (CSIC Iniciación_2017_ID_159) to N. C., Alexander von Humboldt Foundation (AvHF) to T. G. and R. R., the Novo Nordisk Foundation ( NNF13OC0004294 and NNF20SA0064214 ) to M. J. D., and PICT 2018-0795 from Agencia I+d+i Argentina , University of Buenos Aires (grant 20020120300025BA ), and CONICET (grant 11220150100303CO ) to D. E. N. C. was partially supported by a fellowship from Comisión Académica de Posgrado (CAP), Universidad de la República , Uruguay. Additional funding was obtained from Programa de Desarrollo de las Ciencias Básicas (PEDECIBA, Uruguay), Agencia Nacional de Investigación e Innovación (ANII-SNI, Uruguay), EU-LAC Health (EULACH16/T01-0131), and Programa de Alimentos y Salud Humana (PAyS, Uruguay). Funding Information: The authors would like to thank Dr Tobias Karlberg and Dr Susanne Gräslund from the Karolinska Institutet - Structural Genomics Consortium for providing the HsGS plasmid (Construct ID GLULA-c004). We also thank Dr Verónica Tórtora for her major aid within the initial steps of the transformation and HsGS expression experiments. N. C. D. E. P. H. T. G. M. J. D. S. B. and R. R. conceptualization; N. C. Mauricio Mastrogiovanni, Michele Mariotti, F. M. I. and P. H. methodology; N. C. Mauricio Mastrogiovanni, Michele Mariotti, and F. M. I. investigation; N. C. writing–original draft; M. J. D. S. B. and R. R. writing–review and editing; R. R. supervision. This work was supported by grants of Universidad de la República (CSIC_2018 and EI_2020) to R. R. Universidad de la República (CSIC Iniciación_2017_ID_159) to N. C. Alexander von Humboldt Foundation (AvHF) to T. G. and R. R. the Novo Nordisk Foundation (NNF13OC0004294 and NNF20SA0064214) to M. J. D. and PICT 2018-0795 from Agencia I+d+i Argentina, University of Buenos Aires (grant 20020120300025BA), and CONICET (grant 11220150100303CO) to D. E. N. C. was partially supported by a fellowship from Comisión Académica de Posgrado (CAP), Universidad de la República, Uruguay. Additional funding was obtained from Programa de Desarrollo de las Ciencias Básicas (PEDECIBA, Uruguay), Agencia Nacional de Investigación e Innovación (ANII-SNI, Uruguay), EU-LAC Health (EULACH16/T01-0131), and Programa de Alimentos y Salud Humana (PAyS, Uruguay). Publisher Copyright: © 2023 The Authors |
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2023 |
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2023-09-30T22:21:36Z 2023-03 2023-03-01T00:00:00Z |
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