Appearance of a Soluble Form of the G Protein of Respiratory Syncytial Virus in Fluids of Infected Cells

• Published in: Journal of general virology 1987-06, Vol.68 (6), p.1705-1714
• Main Authors: Hendricks, David A, Baradaran, Khandan, McIntosh, Kenneth, Patterson, Jean L
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Capsid - analysis; Cell Extracts - analysis; Cell Line; Culture Media - analysis; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Humans; Membrane Glycoproteins; Microbiology; Molecular Weight; Replicative cycle, interference, host-virus relations, pathogenicity, miscellaneous strains; respiratory syncytial virus; Respiratory Syncytial Viruses - metabolism; Solubility; Viral Core Proteins - analysis; Viral Envelope Proteins; Viral Fusion Proteins - analysis; Viral Proteins - analysis; Viral Proteins - metabolism; Virology
• ISSN: 0022-1317; 1465-2099
• DOI: 10.1099/0022-1317-68-6-1705

1 Division of Infectious Diseases, Department of Pediatrics, The Children's Hospital, 300 Longwood Avenue and 2 Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts 02115, U.S.A. We examined the antigenic reactivities and virion associations of glycoproteins that were released into the culture fluids of cells infected with respiratory syncytial (RS) virus. Culture fluids and cell extracts were obtained from cells 24 to 30 h after they were infected with the Long strain of RS virus. Radioimmune precipitation of [ 3 H]glucosamine-labelled glycoproteins by large glycoprotein (G)-specific or fusion protein (F)-specific monoclonal antibodies (MAbs) revealed that the G, F 1 and F 2 proteins were present in cell extracts but only the G protein was clearly evident in culture fluids. A glycoprotein ( M r 43K) which may be a precursor or a breakdown product of the G protein was also precipitated by the G-specific MAb from cell extracts and culture fluids. The G protein in culture fluids was slightly smaller ( M r 82K) than the G protein in cell extracts ( M r 88K). An abundant or heavily labelled, 18K glycoprotein in the fluids of virus-infected but not of mock-infected cells was weakly precipitated by the F-specific MAb; this suggested that the 18K protein shares epitopes with the fusion protein of RS virus. The absence of F 1 and F 2 polypeptides from culture fluids is evidence that the cells, which contained an abundance of these proteins, were intact. To determine whether any of the viral glycoproteins released by infected cells were soluble (non-virion-associated), culture fluid was subjected to rate zonal centrifugation in a 10 to 50% sucrose gradient. An assay of fractions using a MAb-capture ELISA for the nucleocapsid (N) and F proteins revealed a peak of activity, due to virions, in the centre of the gradient, and a strong signal for the N protein at the top of the gradient suggesting that N protein was released from intact cells. Radioimmune precipitation of glycoproteins from the fractions at the top of the gradient using a hyperimmune guineapig serum revealed the G protein and a heterogeneous band which had the electrophoretic mobility of the 43K protein. Neither the F 1 nor the F 2 protein was present in these fractions thus suggesting that virions had remained intact. These results showed that a soluble form of the G protein of RS virus is released into the culture fluids of intact, infected cells. Several th