Abstract
PCR fragments containing the fusion protein genes 129L of the ectromelia virus (EV) and A30L of the variola virus (VARV) were cloned in pQE32. The expression products, recombinant prA30L and pr129L, were isolated from Escherichia coli cell lysates by metal-chelate affinity chromatography. The recombinant proteins retained the capability of oligomerization, characteristic of their natural analogs. ELISA and immunoblotting were used to test 22 monoclonal antibodies (mAbs) to orthopoxviruses (19 mAbs to EV, 2 mAbs to the vaccinia virus (VACV), and 1 mAb to the cowpox virus (CPXV)) for interaction with prA30L, pr129L, and orthopoxviruses. Twelve species-specific epitopes were found in the EV fusion protein 129L and its recombinant analog. Ten cross-reacting epitopes were found in the EV, CPXV, and VACV fusion proteins. Of these, nine epitopes were present both in prA30L and in the VARV fusion protein. Five mAbs interacting with cross-reacting epitopes were capable of efficient neutralization of VACV; two of these mAbs neutralized VARV. It was demonstrated that there are species-specific epitopes in EV 129L and cross-reacting epitopes in the EV, VARV, CPXV, and VACV fusion proteins, including epitopes that induced synthesis of virus-neutralizing antibodies against VACV and VARV.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
REFERENCES
Marennikova S.S., Shchelkunov S.N. 1998. Patogennye dlya cheloveka ortopoksvirusy (Orthopoxviruses Pathogenic for humans). Moscow: KMK Press.
Moss B. 2000. Poxviridae: The viruses and their repluication. In: Fields in Virology, 4th ed. Eds. Fields B.N., Knipe D.M., Howley P.M., et al. Philadelphia: Lippincott-Raven, 2849–2880.
Moss B. 1991. Vaccinia virus: A tool for research and vaccine development. Science. 252, 1662–1667.
Hutin Y.J., Williams R.J., Malfait P., Pebody R., Loparev V.N., Ropp S.L., Rodriguez M., Knight J.C., Tshioko F.K., Khan A.S., Szczeniowski M.V., Esposito J.J. 2001. Outbreak of human monkeypox, Democratic Republic of Congo, 1996 to 1997. Emerg. Infect. Dis. 7, 434–438.
Guarner J., Johnson B.J., Paddock C.D., Shieh W.J., Goldsmith C.S., Reynolds M.G., Damon I.K., Regnery R.L., Zaki S.R. 2004. Veterinary Monkeypox Virus Working Group. Monkeypox transmission and pathogenesis in prairie dogs. Emerg. Infect. Dis. 10, 426–431.
Gordon J., Mohandas A., Wilton S., Dales S. 1991. A prominent antigenic surface polypeptide involved in the biogenesis and function of the vaccinia virus envelope. Virology. 181, 671–686.
Rodriguez J.F., Esteban M. 1987. Mapping and nucleotide sequence of the vaccinia virus gene that encodes a 14-kilodalton fusion protein. J. Virol. 61, 3550–3554.
Galmiche M.C., Goenaga J., Wittek R., Rindisbacher L. 1999. Neutralizing and protective antibodies directed against vaccinia virus envelope antigens. Virology. 254, 71–80.
Hsiao J.C., Chung C.S., Chang W. 1999. Vaccinia virus envelope D8L protein binds to cell surface chondroitin sulfate and mediates the adsorption of intracellular mature virions to cells. J. Virol. 73, 8750–8761.
Ichihashi Y., Oie M. 1988. Epitope mosaic on the surface proteins of orthopoxviruses. Virology. 163, 133–144.
Wolffe E.J., Vijaya S., Moss B. 1995. A myristylated membrane protein encoded by the vaccinia virus L1R open reading frame is the target of potent neutralizing monoclonal antibodies. Virology. 211, 53–63.
Hooper J.W., Custer D.M., Schmaljohn C.S., Schmaljohn A.L. 2000. DNA vaccination with vaccinia virus L1R and A33R genes protects mice against a lethal poxvirus challenge. Virology. 266, 329–339.
Vazquez M.I., Rivas G., Cregut D., Serrano L., Esteban M. 1998. The vaccinia virus 14-kilodalton (A27L) fusion protein forms a triple coiled-coil structure and interacts with the 21-kilodalton (A17L) virus membrane protein through a C-terminal alpha-helix. J. Virol. 72, 10126–10137.
Wallengren K., Risco C., Krijnse-Locker J., Esteban M., Rodriguez D. 2001. The A17L gene product of vaccinia virus is exposed on the surface of IMV. Virology. 290, 143–152.
Shchelkunov S.N. 1995. Functional organization of variola major and vaccinia virus genomes. Virus Genes. 10, 53–71.
Chen N., Danila M.I., Feng Z., Buller R.M., Wang C., Han X., Lefcovitz E.J., Upton C. 2003. The genomic sequence of ectromelia virus, the causative agent of mousepox. Virology. 317, 168–186
Joklik W.K. 1962. The purification of four strains of poxvirus. Virology. 18, 9–18.
Muravlev A.I., Agafonov A.P., Reshetnikov S.S., Lavrinenko I.A., Cheshenko I.O., Beklemishev A.B. 1995. Protective activity of vaccinia virus envelope proteins isolated using nonionic detergents. Vopr. Virusol. 40, 154–158.
Sambrook J., Fritsch E.F, Maniatis T. 1989. Molecular Cloning: A Laboratory Manual. Cold Spring Harbor, N.Y.: Cold Spring Harbor Lab. Press.
Bradford M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248–254.
Gefter M.L., Margulies D.H., Scharft M. D. 1977. A simple method for polyethylene glycol-promoted hybridization of mouse myeloma cells. Somat. Cell Genet. 3, 231–236.
Razumov I.A., Agapov E.V., Pereboev A.V., Protopopova E.V., Lebedeva S.D., Loktev V.B. 1991. Antigenic structure of the Venezuelan equine encephalomyelitis E2 glycoprotein as analyzed with rat monoclonal antibodies. Vopr. Virusol. 36, 34–37.
Razumov I.A., Vasilieva M.A., Serova O.A., Artemyeva L.V., Belanov E.F., Kochneva G.V., Konovalov E.E., Loktev V.B. 2004. Monoclonal antibodies against ectromelia virus: Neutralizing activity and cross reactivity with orthopoxviruses pathogenic for humans. Vestn. Ross. Acad. Med. Nauk. 8, 19–22.
Bazin H., Malache I.M., Nisol F., Delaunay T. 1990. In: Rat Hybridomas and Rat Monoclonal Antibodies. Ed. Bazin H. Florida: CRC Press, 165–201.
Baker O.R., Bray M., Huggins J.W. 2003. Potential antiviral therapeutics for smallpox, monkeypox and other orthopoxvirus infections. Antiviral Res. 53, 13–23.
Laemmli U.K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 227, 680–685.
Czerny C.P., Johann S., Holzle L., Meyer H. 1994. Epitope detection in the envelope of intracellular naked orthopox viruses and identification of encoding genes. Virology. 200, 764–777.
Meyer H., Osterrieder N., Czerny C.P. 1994. Identification of binding sites for neutralizing monoclonal antibodies on the 14-kDa fusion protein of orthopox viruses. Virology. 200, 778–783.
Dallo S., Rodriguez J.F., Esteban M.A. 1987. 14-kDa envelope protein of vaccinia virus with an important role in virus-host cell interactions is altered during virus persistence and determines the plaque size phenotype of the virus. Virology. 159, 423–432.
Rodriguez J.F., Paez E., Esteban M. 1987. A 14.000-Mr envelope protein of vaccinia virus is involved in cell fusion and forms covalently linked trimers. J. Virol. 61, 395–404.
Va'zques M.-A., Esteban M. 1999. Identification of functional domains in the 14-kilodalton envelope protein (A27L) of vaccinia virus. J. Virol. 73, 9098–9109.
Lin T.-H., Chia C.-M., Hsiao J.-C., Chang W., Ku C.-C., Hung S.-C., Tzou D.-L.M. 2002. Structural analysis of the extracellular domain of vaccinia virus envelope protein, A27L, by NMR and CD spectroscopy. J. Biol. Chem. 277, 20949–20959.
Rodriguez J.F., Janeczko R., Esteban M. 1985. Isolation and characterization of neutralizing monoclonal antibodies to vaccinia virus. J. Virol. 56, 482–488.
Author information
Authors and Affiliations
Additional information
__________
Translated from Molekulyarnaya Biologiya, Vol. 39, No. 6, 2005, pp. 1046–1054.
Original Russian Text Copyright © 2005 by Razumov, Gileva, Vasil'eva, Nepomnyashchikh, Mishina, Belanov, Kochneva, Konovalov, Shchelkunov, Loktev.
Rights and permissions
About this article
Cite this article
Razumov, I.A., Gileva, I.P., Vasil'eva, M.A. et al. Neutralizing Monoclonal Antibodies Cross-React with Fusion Proteins Encoded by 129L of the Ectromelia Virus and A30L of the Variola Virus. Mol Biol 39, 918–925 (2005). https://doi.org/10.1007/s11008-005-0113-x
Received:
Issue Date:
DOI: https://doi.org/10.1007/s11008-005-0113-x