| Author | Allonso, Diego | |
| Author | Pereira, Iuri B. | |
| Author | Alves, Ada M. B. | |
| Author | Kurtenbach, Eleonora | |
| Author | Mohana-Borges, Ronaldo | |
| Access date | 2019-09-03T13:12:18Z | |
| Available date | 2019-09-03T13:12:18Z | |
| Document date | 2019 | |
| Citation | ALLONSO, Diego et al. Expression of soluble, glycosylated and correctly folded dengue virus NS1 protein in Pichia pastoris. Protein Expression and Purification, v. 162, p. 9-17, May 2019. | pt_BR |
| ISSN | 1046-5928 | pt_BR |
| URI | https://www.arca.fiocruz.br/handle/icict/35298 | |
| Language | eng | pt_BR |
| Publisher | Elsevier | pt_BR |
| Rights | restricted access | |
| Subject in Portuguese | Vírus da Dengue | pt_BR |
| Subject in Portuguese | Glicosilação | pt_BR |
| Subject in Portuguese | Pichia pastoris | pt_BR |
| Subject in Portuguese | Proteína NS1 | pt_BR |
| Title | Expression of soluble, glycosylated and correctly folded dengue virus NS1 protein in Pichia pastoris | pt_BR |
| Type | Article | |
| DOI | 10.1016/j.pep.2019.05.003 | |
| Abstract | The dengue virus (DENV) non structural protein 1 (NS1) is a 46-55 kDa protein that exists as homodimer inside cells and as hexamer in the extracellular milieu. Several lines of evidence have demonstrated that the biochemical and structural properties of recombinant NS1 (rNS1) vary depending on the protein expression system used. Aiming to improve current tools for studying NS1 multiple roles, a recombinant tag-free NS1 protein was expressed and purified from P. pastoris yeast cells. The best expression condition was achieved using GS115 strain and induction for 72 h with 0.7% methanol addition every 24 h. Total secreted rNS1 reached 2.18 mg in 1 L culture and the final yield of the purified protein was 1 mg per liter of culture (recovery yield of approximately 45.9%). Size-exclusion chromatography and treatment with EndoH and PNGase indicate that it exists as an N-glycosylated homodimer. Moreover, an ELISA assay with specific DENV anti-NS1 antibody that recognizes conformational epitopes revealed that rNS1 has most of its conformational epitopes preserved. The expression of rNS1 in its native conformation is an important tool for further studies of its role in DENV pathogenesis and replication. | pt_BR |
| Affilliation | Universidade Federal do Rio de Janeiro. Faculdade de Farmácia. Departamento de Biotecnologia Farmacêutica. Rio de Janeiro, RJ, Brasil / Universidade Federal do Rio de Janeiro. Instituto de Biofísica Carlos Chagas Filho. Laboratório de Biotecnologia e Bioengenharia Estrutural. Rio de Janeiro, RJ, Brasil. | pt_BR |
| Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Biofísica Carlos Chagas Filho. Laboratório de Bioquímica e Biologia Molecular de Proteínas. Rio de Janeiro, RJ, Brasil. | pt_BR |
| Affilliation | Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biotecnologia e Fisiologia de Infecções Virais. Rio de Janeiro, RJ, Brasil. | pt_BR |
| Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Biofísica Carlos Chagas Filho. Laboratório de Bioquímica e Biologia Molecular de Proteínas. Rio de Janeiro, RJ, Brasil. | pt_BR |
| Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Biofísica Carlos Chagas Filho. Laboratório de Biotecnologia e Bioengenharia Estrutural. Rio de Janeiro, RJ, Brasil | pt_BR |
| Subject | Dengue virus | pt_BR |
| Subject | NS1 protein | pt_BR |
| Subject | Pichia pastoris | pt_BR |
| Subject | Glycosylation | pt_BR |
| DeCS | Glicosilação | pt_BR |
| e-ISSN | 1096-0279 | |
| Embargo date | 2025-01-01 | |
