title: Production of a Recombinant Dengue Virus 2 NS5 Protein and Potential Use as a Vaccine Antigen.
abstract: Dengue fever is caused by any of the four known dengue virus serotypes (DENV1 to DENV4) that affect millions of people worldwide, causing a significant number of deaths. There are vaccines based on chimeric viruses, but they still are not in clinical use. Anti-DENV vaccine strategies based on nonstructural proteins are promising alternatives to those based on whole virus or structural proteins. The DENV nonstructural protein 5 (NS5) is the main target of anti-DENV T cell-based immune responses in humans. In this study, we purified a soluble recombinant form of DENV2 NS5 expressed in Escherichia coli at large amounts and high purity after optimization of expression conditions and purification steps. The purified DENV2 NS5 was recognized by serum from DENV1-, DENV2-, DENV3-, or DENV4-infected patients in an epitope-conformation-dependent manner. In addition, immunization of BALB/c mice with NS5 induced high levels of NS5-specific antibodies and expansion of gamma interferon- and tumor necrosis factor alpha-producing T cells. Moreover, mice immunized with purified NS5 were partially protected from lethal challenges with the DENV2 NGC strain and with a clinical isolate (JHA1). These results indicate that the recombinant NS5 protein preserves immunological determinants of the native protein and is a promising vaccine antigen capable of inducing protective immune responses.
authors.affiliation: Laboratório de Desenvolvimento de Vacinas, Instituto de Ciências Biomédicas, Departamento de Microbiologia, Universidade de São Paulo, São Paulo, Brazil.
authors.forename: Rúbens Prince Dos Santos
authors.initials: RPDS
authors.lastname: Alves
authors.affiliation: Laboratório de Desenvolvimento de Vacinas, Instituto de Ciências Biomédicas, Departamento de Microbiologia, Universidade de São Paulo, São Paulo, Brazil.
authors.forename: Lennon Ramos
authors.initials: LR
authors.lastname: Pereira
authors.affiliation: Laboratório de Desenvolvimento de Vacinas, Instituto de Ciências Biomédicas, Departamento de Microbiologia, Universidade de São Paulo, São Paulo, Brazil.
authors.forename: Denicar Lina Nascimento
authors.initials: DLN
authors.lastname: Fabris
authors.affiliation: Instituto de Medicina Tropical, Escola de Medicina, Universidade de São Paulo, São Paulo, Brazil.
authors.forename: Felipe Scassi
authors.initials: FS
authors.lastname: Salvador
authors.affiliation: Laboratório de Desenvolvimento de Vacinas, Instituto de Ciências Biomédicas, Departamento de Microbiologia, Universidade de São Paulo, São Paulo, Brazil.
authors.forename: Robert Andreata
authors.initials: RA
authors.lastname: Santos
authors.affiliation: Laboratório de Evolução Molecular e Bioinformática, Instituto de Ciências Biomédicas, Departamento de Microbiologia, Universidade de São Paulo, São Paulo, Brazil.
authors.forename: Paolo Marinho de Andrade
authors.initials: PMA
authors.lastname: Zanotto
authors.affiliation: Instituto de Medicina Tropical, Escola de Medicina, Universidade de São Paulo, São Paulo, Brazil.
authors.forename: Camila Malta
authors.initials: CM
authors.lastname: Romano
authors.affiliation: Laboratório de Desenvolvimento de Vacinas, Instituto de Ciências Biomédicas, Departamento de Microbiologia, Universidade de São Paulo, São Paulo, Brazil jaime.amorim@ufob.edu.br lcsf@usp.br.
authors.forename: Jaime Henrique
authors.initials: JH
authors.lastname: Amorim
authors.affiliation: Laboratório de Desenvolvimento de Vacinas, Instituto de Ciências Biomédicas, Departamento de Microbiologia, Universidade de São Paulo, São Paulo, Brazil jaime.amorim@ufob.edu.br lcsf@usp.br.
authors.forename: Luís Carlos de Souza
authors.initials: LCS
authors.lastname: Ferreira
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D003198:Computer Simulation; D003715:Dengue / Q000276:immunology / Q000517:prevention & control* / Q000821:virology; D053059:Dengue Vaccines / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D003716:Dengue Virus / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D000939:Epitopes / Q000032:analysis / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007111:Immunity, Cellular; D051379:Mice; D008807:Mice, Inbred BALB C; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D017361:Viral Nonstructural Proteins / Q000008:administration & dosage / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification
chemical_list: D000914:Antibodies, Viral; D053059:Dengue Vaccines; D000939:Epitopes; C582223:NS5 protein, dengue virus; D011994:Recombinant Proteins; D017361:Viral Nonstructural Proteins