Isolation of the dengue virus envelope glycoprotein from membranes of infected cells by concanavalin A affinity chromatography.
The membranes isolated from type 2 dengue virus-infected BHK-21/15 cells contain three glycosylated virus-specified proteins; one corresponds to the virion envelope glycoprotein, V-3, whereas the other two are nonstructural virus-specified proteins, NV-2 and NV-3. A combination of two nonionic detergents, Nonidet P-40 and Triton X-305, solubilized greater than or equal to 80% of the membrane-bound protein and the majority of the type 2 dengue virus complement-fixing antigens. The soluble material was adsorbed by concanavalin A-Sepharose in the presence of the nonionic detergents, which were subsequently removed by washing with deoxycholate-containing buffer. Finally, the bound glycoprotein was eluted by the addition of alpha-methyl glucopyranoside. V-3 was the only virus-specified protein in the alpha-methyl glucopyranoside eluate. The V-3-containing fraction did not cross-react with antisera against other selected Flaviviruses in the complement fixation tests. The V-3 contained in the isolated fraction differed from the parent membrane-bound V-3 in two interesting, and as yet unexplained, ways: (i) it exhibited hemagglutinating activity similar to that of the infectious virus, but (ii) it did not block the action of neutralizing antibody.
D000956:Antigens, Viral; D002460:Cell Line; D002462:Cell Membrane / Q000032:analysis; D002846:Chromatography, Affinity; D003716:Dengue Virus / Q000032:analysis* / Q000276:immunology; D006023:Glycoproteins / Q000276:immunology / Q000302:isolation & purification*; D008757:Methylglucosides; D012997:Solvents; D013501:Surface-Active Agents; D014764:Viral Proteins / Q000276:immunology / Q000302:isolation & purification*
D000956:Antigens, Viral; D006023:Glycoproteins; D008757:Methylglucosides; D012997:Solvents; D013501:Surface-Active Agents; D014764:Viral Proteins