Assessment of antigenicity and genetic variation of glycoprotein B of murine cytomegalovirus.
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 of sequence heterogeneity consistent with these findings.
Xu
J
J
Department of Microbiology, University of Western Australia, Nedlands, Australia.
Lyons
P A
PA
Carter
M D
MD
Booth
T W
TW
Davis-Poynter
N J
NJ
Shellam
G R
GR
Scalzo
A A
AA
D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology; D000956:Antigens, Viral / Q000276:immunology; D001483:Base Sequence; D001665:Binding Sites; D001666:Binding Sites, Antibody / Q000235:genetics; D002460:Cell Line; D017931:DNA Primers; D004926:Escherichia coli; D005260:Female; D014644:Genetic Variation; D006566:Herpesviridae Infections / Q000517:prevention & control; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D018146:Muromegalovirus / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification; D009500:Neutralization Tests; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D017386:Sequence Homology, Amino Acid; D014611:Vaccination; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000276:immunology*
D000914:Antibodies, Viral; D000956:Antigens, Viral; D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C040495:glycoprotein B, Simplexvirus