title: Dengue-1 envelope protein domain III produced in Pichia pastoris: potential use for serological diagnosis.
abstract: Dengue is a major international public health concern. There is no drug to treat dengue virus infections and a vaccine is yet to be licensed. The laboratory diagnosis of dengue virus infection has been greatly improved during the last decade; therefore, the main limiting factor is the production of recombinant viral antigens on a large scale. Domain III of dengue virus envelope protein contains multiplex conformation-dependent neutralizing epitopes, making it an attractive diagnostic candidate. In this work, we have demonstrated the expression of dengue virus type 1 envelope domain III protein (EDIII-D1) in methylotrophic yeast, Pichia pastoris GS115. The recombinant secreted protein (sEDIII-D1) was purified by affinity chromatography and characterized by SDS-PAGE. Purified protein was recognized in immunoblot analysis and enzyme-linked immunosorbent assay (ELISA) with dengue-infected human serum samples. In conclusion, secreted expressions of domain III protein can be obtained in P. pastoris by methanol induction. This product has the potential to be used for the diagnosis of dengue infections.
authors.affiliation: Laboratory of Molecular Immunovirology, Department of General Biology, Federal University of Viçosa, Viçosa (MG), Brazil.
authors.forename: Silvia A
authors.initials: SA
authors.lastname: Cardoso
authors.forename: Vinícus F
authors.initials: VF
authors.lastname: Paixão
authors.forename: Michelle D
authors.initials: MD
authors.lastname: Oliveira
authors.forename: Eduardo R
authors.initials: ER
authors.lastname: Honda
authors.forename: Leandro L
authors.initials: LL
authors.lastname: Oliveira
authors.forename: Cynthia C
authors.initials: CC
authors.lastname: da Silva
authors.forename: Sérgio O
authors.initials: SO
authors.lastname: De Paula
mesh_terms: D000595:Amino Acid Sequence; D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology; D000956:Antigens, Viral / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D003715:Dengue / Q000097:blood / Q000175:diagnosis* / Q000276:immunology / Q000821:virology*; D003716:Dengue Virus / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood / Q000276:immunology; D007075:Immunoglobulin M / Q000097:blood / Q000276:immunology; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins
keywords: Dengue; Diagnosis; Domain III; ELISA; Expression; Pichia pastoris