Assessment of antigenicity and genetic variation of glycoprotein B of murine cytomegalovirus

• Published in: Journal of general virology 1996-01, Vol.77 (1), p.49-59
• Main Authors: Xu, Jiake, Lyons, Paul A, Carter, Meredith D, Booth, Timothy W. M, Davis-Poynter, Nicholas J, Shellam, Geoffrey R, Scalzo, Anthony A
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Antibodies, Viral - immunology; Antigens, Viral - immunology; Base Sequence; Binding Sites; Binding Sites, Antibody - genetics; Cell Line; DNA Primers; Escherichia coli; Female; Genetic Variation; Herpesviridae Infections - prevention & control; Mice; Mice, Inbred BALB C; Molecular Sequence Data; murine cytomegalovirus; Muromegalovirus - genetics; Muromegalovirus - immunology; Muromegalovirus - isolation & purification; Neutralization Tests; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - immunology; Sequence Homology, Amino Acid; Vaccination; Viral Envelope Proteins - genetics; Viral Envelope Proteins - immunology
• ISSN: 0022-1317; 1465-2099
• DOI: 10.1099/0022-1317-77-1-49

Department of Microbiology, University of Western Australia, Nedlands, 6907 Western Australia, Australia An analysis of linear antibody-binding sites of the glycoprotein B (gB) molecule of murine cytomegalovirus (MCMV) and of genetic variation within these regions was performed. To achieve this, a series of overlapping fragments spanning the entire coding sequence of the gB gene of the K181 strain of MCMV was expressed in E. coli as fusion proteins with glutathione S -transferase (GST) using the pGEX expression system. Four antibody-binding regions were mapped to locations spanning amino acid residues 17–79 (BS), 155–278 (BE2), 809–926 (SS) and 347–508 (BB and EE), based on reactivity in Western blot analysis of GST-gB fusion proteins with murine polyclonal antiserum raised against MCMV. Only the antibody-binding region BE2 (155–278) elicited an antiserum that exhibited complement-dependent neutralizing activity, and immunization of mice with the fusion protein BE2 led to moderate but significant reductions in the level of MCMV replication in the spleen. Polyclonal antisera raised against the GST-gB fusion proteins detected purified virion proteins of 105 kDa (anti-BS and anti-BE2) and 52 kDa (anti-SS), and are therefore likely to recognize the N-terminal and C-terminal portions of the gB molecule, respectively. The antibody-binding region within amino acid residues 17–79 was found to be MCMV strain-specific, whereas antibody-binding regions within residues 155–278 and 809–926 were found to be conserved among MCMV field isolates. Comparative sequence analysis of the corresponding regions of MCMV gB revealed a level and extent of sequence heterogeneity consistent with these findings. * Author for correspondence. Fax [phone]. e-mail [e-mail] Present address: Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305-5402, USA. Present address: Wellcome Trust Centre for Human Genetics, Nuffield Orthopaedic Centre, University of Oxford, Headington, Oxford OX3 7BN, UK. Present address: Institute for Child Health Research, Subiaco, 6008 Western Australia, Australia. The nucleotide and deduced amino acid sequences reported in this paper have been submitted to the Genome Sequence Data Base and assigned the accession numbers L39243 [GenBank] -L39251, L39215 [GenBank] -L39228, L39252 [GenBank] -L39260 and L39229 [GenBank] -L39242. Received 13 June 1995; accepted 7 September 1995.