title: Purified dengue 2 virus envelope glycoprotein aggregates produced by baculovirus are immunogenic in mice.
abstract: The full-length dengue 2 virus envelope glycoprotein (Egp) was expressed in insect cells by recombinant (r) baculovirus and found to form multimeric aggregates that were recovered in the void volume of gel filtration columns and by ultracentrifugation. An immunoblot confirmed that rEgp aggregrates disrupted with SDS sample buffer released a monomeric form that migrated with a molecular weight similar to native dengue 2 virus Egp on polyacrylamide gels. The rEgp aggregates reacted strongly with a panel of monoclonal antibodies specific for the native Egp and which identify critical structural and functional epitopes. The rEgp aggregates were purified by ultracentrifugation through 30% sucrose, and were shown to be the major protein band on a polyacrylamide gel and corresponding immunoblot. Purified rEgp aggregates in combination with aluminum hydroxide induced high titer neutralizing antibodies in adult mice. The generation of full-length dengue 2 rEgp aggregates in insect cells facilitated development of a simple, effective procedure for purification of the recombinant protein, and represents a good approach for producing highly immunogenic dengue 2 rEgp as a component of a subunit vaccine.
authors.affiliation: Department of Virus Diseases, Walter Reed Army Institute of Research, Washington, DC 20307, USA. eileen.kelly@na.amedd.army.mil
authors.forename: E P
authors.initials: EP
authors.lastname: Kelly
authors.forename: J J
authors.initials: JJ
authors.lastname: Greene
authors.forename: A D
authors.initials: AD
authors.lastname: King
authors.forename: B L
authors.initials: BL
authors.lastname: Innis
mesh_terms: D000276:Adjuvants, Immunologic; D000536:Aluminum Hydroxide; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000914:Antibodies, Viral / Q000096:biosynthesis* / Q000276:immunology; D002460:Cell Line; D002850:Chromatography, Gel; D003716:Dengue Virus / Q000235:genetics / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel; D005069:Evaluation Studies as Topic; D005260:Female; D005822:Genetic Vectors / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D009036:Moths / Q000166:cytology; D017924:Nucleopolyhedroviruses / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification; D018411:Spodoptera / Q000166:cytology; D014611:Vaccination; D014614:Vaccines, Synthetic / Q000276:immunology*; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D010948:Viral Plaque Assay
chemical_list: D000276:Adjuvants, Immunologic; D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; C051821:E-glycoprotein, Dengue virus type 2; D011993:Recombinant Fusion Proteins; D014614:Vaccines, Synthetic; D014759:Viral Envelope Proteins; D000536:Aluminum Hydroxide