title: Pichia pastoris-expressed dengue virus type 2 envelope domain III elicits virus-neutralizing antibodies.
abstract: A tetravalent dengue vaccine that can protect against all four serotypes of dengue viruses is a global priority. The host-receptor binding, multiple neutralizing epitope-containing carboxy-terminal region of the dengue envelope protein, known as domain III (EDIII), has emerged as a promising subunit vaccine antigen. One strategy to develop a tetravalent dengue subunit vaccine envisages mixing recombinant EDIIIs, corresponding to the four dengue virus serotypes. Towards this objective, a recombinant clone of the methylotrophic yeast Pichia pastoris, harboring the EDIII gene of dengue virus type 2 (EDIII-2) for its intracellular expression, was created. Recombinant EDIII-2 protein, expressed by this clone was purified to near homogeneity by affinity chromatography, with final yields of >50mg/l culture. Groups of Balb/c mice were immunized with this protein, separately formulated in two adjuvants, alum and montanide ISA 720. The EDIII-2 antigen, formulated in either adjuvant, elicited high levels of neutralizing antibodies to dengue virus type 2 in mice as analyzed by Plaque Reduction Neutralization Test (PRNT). This study demonstrates the feasibility of using P. pastoris to produce EDIII antigens capable of eliciting potent virus-neutralizing antibodies.
authors.affiliation: Recombinant Gene Products Group, International Centre for Genetic Engineering & Biotechnology, New Delhi, India.
authors.forename: Gaurav
authors.initials: G
authors.lastname: Batra
authors.forename: Rajendra
authors.initials: R
authors.lastname: Raut
authors.forename: Satinder
authors.initials: S
authors.lastname: Dahiya
authors.forename: Neha
authors.initials: N
authors.lastname: Kamran
authors.forename: Sathyamangalam
authors.initials: S
authors.lastname: Swaminathan
authors.forename: Navin
authors.initials: N
authors.lastname: Khanna
mesh_terms: D000276:Adjuvants, Immunologic / Q000008:administration & dosage; D000534:Alum Compounds / Q000008:administration & dosage; D000818:Animals; D057134:Antibodies, Neutralizing / Q000097:blood*; D000914:Antibodies, Viral / Q000097:blood*; D053059:Dengue Vaccines / Q000235:genetics / Q000276:immunology*; D003716:Dengue Virus / Q000276:immunology; D008353:Mannitol / Q000008:administration & dosage / Q000031:analogs & derivatives; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D009829:Oleic Acids / Q000008:administration & dosage; D010843:Pichia / Q000235:genetics; D017422:Sequence Analysis, DNA; D022223:Vaccines, Subunit / Q000235:genetics / Q000276:immunology; D014614:Vaccines, Synthetic / Q000235:genetics / Q000276:immunology; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology*; D010948:Viral Plaque Assay
chemical_list: D000276:Adjuvants, Immunologic; D000534:Alum Compounds; D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D053059:Dengue Vaccines; C051821:E-glycoprotein, Dengue virus type 2; D009829:Oleic Acids; D022223:Vaccines, Subunit; D014614:Vaccines, Synthetic; D014759:Viral Envelope Proteins; C001681:mannide monooleate; C041524:aluminum sulfate; D008353:Mannitol