Epitope mapping and characterization of the infectious hematopoietic necrosis virus glycoprotein, using fusion proteins synthesized in Escherichia coli.
A characterization of the antigenic determinants (epitopes) of the glycoprotein (G) of infectious hematopoietic necrosis virus was made by expressing different regions of the G gene in Escherichia coli. A cDNA copy of the G gene was divided into four fragments by TaqI digestion, and the fragments were subcloned into pATH vectors, placing the expression of each G gene fragment under control of the trpE promoter. The resulting plasmids, pXL2, pXL3, and pXL7, encoded trpE-G fusion proteins subsequently detected with anti-infectious hematopoietic necrosis virus sera by Western immunoblots. A comparison of reactivities of the fusion proteins encoded by these plasmids was made by Western immunoblot and radioimmunoassay with a number of anti-G specific monoclonal antibodies (MAbs). The nonneutralizing MAb 136J reacted with the trpE-G fusion protein encoded by pXL3 and fusion proteins encoded by plasmids p52G and p618G, which were described in previous studies (R. D. Gilmore, Jr., H. M. Engelking, D. S. Manning, and J. C. Leong. Bio/Technology 6:295-300, 1988). Another nonneutralizing MAb, 2F, bound to the pXL3 fusion protein, and the neutralizing MAb RB/B5 recognized the pXL7 fusion protein. All fusion proteins were tested as vaccines in rainbow trout fry. Although significant protection was induced by all fusion proteins, the pXL3 fusion protein was most effective as a vaccine.
Xu
L
L
Department of Microbiology, Oregon State University, Corvallis 97331-3804.
Mourich
D V
DV
Engelking
H M
HM
Ristow
S
S
Arnzen
J
J
Leong
J C
JC
D000911:Antibodies, Monoclonal; D001483:Base Sequence; D000939:Epitopes / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D005814:Genes, Viral; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000276:immunology; D015183:Restriction Mapping; D012209:Rhabdoviridae / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D014764:Viral Proteins / Q000235:genetics* / Q000276:immunology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D046911:Macromolecular Substances; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins