Summary.
Infection of cells with classical swine fever virus (CSFV) is mediated by the interaction of envelope glycoproteins E2 and Erns with receptor molecules on the cell surface. These proteins are also the major antigens for eliciting neutralizing antibodies and conferring protective immunity. Here we report the identification of multiple neutralizing epitopes on these proteins by screening a phage-displayed random peptide library with CSFV-specific neutralizing monoclonal antibodies. Two different E2-specific neutralizing mAbs (a18 and 24/10) were found to bind to a common motif SPTxL, which is similar to the sequence SPTTL of the E2 protein (aa 289–293), indicating that this is likely to be an immunodominant epitope. Similarly, an immunodominant epitope corresponding to the sequence DKN of Erns (aa 117–119) was identified for two independent Erns-specific neutralizing antibodies, b4-22 and 24/16, respectively. Another binding motif, CxNNxTC, was identified for mAb 24/16, but not for b4-22. Sequencing analysis of the genes coding for the light chain of these mAbs was conducted to ensure that all mAbs were derived from different hybridomas, rather than from different subclones of a common parent line. Inhibition studies using immunofluorescent antibody assay and virus neutralization test demonstrated that the mimotope peptides truly mimicked the antibody binding determinants on the viral proteins. The detailed mapping data for these neutralizing epitopes will be useful for development of improved diagnostic tests and perhaps a peptide-based vaccine for this important swine disease.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
CG Adda RF Anders L Tilley M Foley (2002) ArticleTitleRandom sequence libraries displayed on phage: identification of biologically important molecules. Comb Chem High Throughput Screen 5 1–14 Occurrence Handle11860335 Occurrence Handle1:CAS:528:DC%2BD38XitVWnsL4%3D
M Arnal D Fernandez-de-Luco L Riba M Maley J Gilray K Willoughby S Vilcek PF Nettleton (2004) ArticleTitleA novel pestivirus associated with deaths in Pyrenean chamois (Rupicapra pyrenaica pyrenaica). J Gen Virol 85 3653–3657 Occurrence Handle15557238 Occurrence Handle10.1099/vir.0.80235-0 Occurrence Handle1:CAS:528:DC%2BD2cXhtVOmsbvI
P Becher RA Ramirez M Orlich SC Rosales M König M Schweizer H Stalder H Schirrmeier HJ Thiel (2003) ArticleTitleGenetic and antigenic characterization of novel pestivirus genotypes: implications for classification. Virology 311 96–104 Occurrence Handle12832207 Occurrence Handle10.1016/S0042-6822(03)00192-2 Occurrence Handle1:CAS:528:DC%2BD3sXkvVCnt7g%3D
EA Carbrey WC Stewart JL Kresse M Snijder (1976) ArticleTitleNatural infection of pigs with bovine diarrhoea virus and its differential diagnosis from hog cholera. J Am Vet Med Assoc 169 1217–1219 Occurrence Handle187565 Occurrence Handle1:STN:280:CSiD2s7htlI%3D
A Christmann A Wentzel C Meyer G Meyers H Kolmar (2001) ArticleTitleEpitope mapping and affinity purification of monospecific antibodies by Escherichia coli cell surface display of gene-derived random peptide libraries. J Immunol Methods 257 163–173 Occurrence Handle11687250 Occurrence Handle10.1016/S0022-1759(01)00461-6 Occurrence Handle1:CAS:528:DC%2BD3MXnvFKjsb8%3D
S Edwards V Moennig G Wensvoort (1991) ArticleTitleThe development of an international reference panel of monoclonal antibodies for the differentiation of hog cholera virus from other pestiviruses. Vet Microbiol 29 101–108 Occurrence Handle1660637 Occurrence Handle10.1016/0378-1135(91)90118-Y Occurrence Handle1:STN:280:By2D1c7itFU%3D
RIB Francki CM Fauquet DL Knudson F Brown (1991) ArticleTitleFlaviviridae. Arch Virol [Suppl] 2 223–233
MM Hulst G Himes E Newbigin RJM Moormann (1994) ArticleTitleGlycoprotein E2 of classical swine fever virus: expression in insect cells and identification as a ribonuclease. Virology 200 558–565 Occurrence Handle8178442 Occurrence Handle10.1006/viro.1994.1218 Occurrence Handle1:CAS:528:DyaK2cXktVais7k%3D
MM Hulst RJM Moormann (1997) ArticleTitleInhibition of pestivirus infection in cell culture by envelope proteins Erns and E2 of classical swine fever virus: Erns and E2 interact with different receptors. J Gen Virol 78 2779–2787 Occurrence Handle9367363 Occurrence Handle1:CAS:528:DyaK2sXntVyhs7c%3D
MM Hulst HGP Van Gennip RJM Moormann (2000) ArticleTitlePassage of classical swine fever virus in cultured swine kidney cells selects virus variants that bind to heparan sulfate due to a single amino acid change in envelope protein Erns. J Virol 74 9553–9561 Occurrence Handle11000226 Occurrence Handle10.1128/JVI.74.20.9553-9561.2000 Occurrence Handle1:CAS:528:DC%2BD3cXnt1alsr8%3D
MB Irving O Pan JK Scott (2001) ArticleTitleRandom-peptide libraries and antigen-fragment libraries for epitope mapping and the development of vaccines and diagnostics. Curr Opin Chem Biol 5 314–324 Occurrence Handle11479124 Occurrence Handle10.1016/S1367-5931(00)00208-8 Occurrence Handle1:CAS:528:DC%2BD3MXktlShtbw%3D
A Kosmidou R Ahl HJ Thiel E Weiland (1995) ArticleTitleDifferentiation of classical swine fever virus (CSFV) strains using monoclonal antibodies against structural glycoproteins. Vet Microbiol 47 111–118 Occurrence Handle8604543 Occurrence Handle10.1016/0378-1135(95)00054-E Occurrence Handle1:CAS:528:DyaK28Xjt1Sksg%3D%3D
M König T Lengsfeld T Pauly R Stark HJ Thiel (1995) ArticleTitleClassical swine fever virus: independent induction of protective immunity by two structural proteins. J Virol 69 6479–6486 Occurrence Handle7666549
UK Laemmli (1970) ArticleTitleCleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227 680–685 Occurrence Handle5432063 Occurrence Handle10.1038/227680a0 Occurrence Handle1:CAS:528:DC%2BD3MXlsFags7s%3D
M Lin F Lin M Mallory A Clavijo (2000) ArticleTitleDeletions of structural glycoprotein E2 of classical swine fever virus strain Alfort/187 resolve a linear epitope of monoclonal antibody WH303 and the minimal N-terminal domain essential for binding immunoglobulin G antibodies of a pig hyperimmune serum. J Virol 74 11619–11625 Occurrence Handle11090160 Occurrence Handle1:CAS:528:DC%2BD3MXitVGju7c%3D
Lindenbach BD, Rice CM (2001) Flaviviridae: the viruses and their replication. In: Knipe DM, Howley PM, Griffin DE, Lamb RA, Martin MA, Roizman B, Straus SE (eds) Fields virology, 4th edn. Lippincott-Raven, Philadelphia, pp 991–1041
BT Messmer JJ Sullivan N Chiorazzi TC Rodman DS Thaler (1999) ArticleTitleTwo human neonatal IgM antibodies encoded by different variable-region genes bind the same linear peptide: evidence for a stereotyped repertoire of epitope recognition. J Immunol 162 2184–2192 Occurrence Handle9973494 Occurrence Handle1:CAS:528:DyaK1MXhtVOmtr0%3D
V Moennig J Plagemann (1992) ArticleTitleThe pestiviruses. Adv Virus Res 41 53–91 Occurrence Handle1315479 Occurrence Handle1:CAS:528:DyaK3sXjsVGh
G Meyers H-J Thiel (1996) ArticleTitleMolecular characterization of pestiviruses. Adv Virus Res 47 53–118 Occurrence Handle8895831 Occurrence Handle1:CAS:528:DyaK2sXivVOksg%3D%3D
DJ Rodi L Makowski (1999) ArticleTitlePhage-display technology-finding a needle in a vast molecular haystack. Curr Opin Biotechnol 10 87–93 Occurrence Handle10047512 Occurrence Handle10.1016/S0958-1669(99)80016-0 Occurrence Handle1:CAS:528:DyaK1MXht1Cjtrk%3D
MJ Rowley M Scealy JC Whisstock JA Jois LC Wijeyewickrema IR Mackay (2000) ArticleTitlePrediction of the immunodominant epitope of the pyruvate dehydrogenase complex E2 in primary biliary cirrhosis using phage display. J Immunol 164 3413–3419 Occurrence Handle10706737 Occurrence Handle1:CAS:528:DC%2BD3cXhvVektr8%3D
T Rümenapf G Unger R Stark HJ Thiel (1991) ArticleTitleMolecualr characterization of hog cholera virus. Arch Virol [Suppl] 3 7–18
T Rümenapf G Unger JH Strauss HJ Thiel (1993) ArticleTitleProcessing of the envelope glycoproteins of pestiviruses. J Virol 67 3288–3294 Occurrence Handle8388499
H Schirrmeier G Strebelow K Depner G Hoffmann M Beer (2004) ArticleTitleGenetic and antigenic characterization of an atypical pestivirus isolate, a putative member of a novel pestivirus species. J Gen Virol 85 3647–3652 Occurrence Handle15557237 Occurrence Handle10.1099/vir.0.80238-0 Occurrence Handle1:CAS:528:DC%2BD2cXhtVOmsbvO
R Schneider G Unger R Stark E Schneider-Scherzer HJ Thiel (1993) ArticleTitleIdentification of a structural glycoprotein of an RNA virus as a ribonuclease. Science 261 1169–1171 Occurrence Handle8356450 Occurrence Handle1:CAS:528:DyaK3sXlslKns7Y%3D
C Terpstra M Bloemraad AJL Gielkens (1984) ArticleTitleThe neutralizing peroxidase linked assay for detection of antibody against swine fever virus. Vet Microbiol 9 113–120 Occurrence Handle6375112 Occurrence Handle10.1016/0378-1135(84)90026-9 Occurrence Handle1:STN:280:BiuB38rnsVA%3D
HJ Thiel R Stark E Weiland T Rumenapf G Meyers (1991) ArticleTitleHog cholera virus: molecular composition of virions from a pestivirus. J Virol 65 4705–4712 Occurrence Handle1870198 Occurrence Handle1:CAS:528:DyaK3MXmsFWkt7g%3D
van Regenmortel MHV, Fauquet CM, Bishop DHL, Carstens EB, Estes MK, Lemon SM, Maniloff J, Mayo MA, McGeoch DJ, Pringle CR, Wickner RB (2000) Virus taxonomy. Classification and nomenclature of viruses. Seventh Report of the International Committee on Taxonomy of Viruses. Academic Press, San Diego
PA Van Rijn RGP van Gennip EJ de Meijer RJM Moormann (1992) ArticleTitleA preliminary map of epitopes on envelope glycoprotein E1 of HCV strain Brescia. Vet Microbiol 33 221–230 Occurrence Handle1282755 Occurrence Handle1:CAS:528:DyaK3sXhsVGnsL8%3D
PA Van Rijn HGP van Gennip EJ de Meijer RJM Moormann (1993) ArticleTitleEpitope mapping of envelope glycoprotein E1 of hog cholera virus strain Brescia. J Gen Virol 74 2053–2060 Occurrence Handle7691986 Occurrence Handle1:CAS:528:DyaK3sXms1yqtLw%3D Occurrence Handle10.1099/0022-1317-74-10-2053
PA Van Rijn GKW Miedema G Wensvoort HGP van Gennip RJM Moormann (1994) ArticleTitleAntigenic structure of envelope glycoprotein E1 of hog cholera virus. J Virol 68 3934–3942 Occurrence Handle7514680 Occurrence Handle1:CAS:528:DyaK2cXktlalu7k%3D
L-F Wang M Yu (2004) ArticleTitleEpitope identification using phage display libraries. Curr Drug Targets 5 1–15 Occurrence Handle14738215 Occurrence Handle10.2174/1389450043490668
L-F Wang M Yu J White BT Eaton (1996) ArticleTitleBTag: a novel six-residue epitope tag for surveillance and purification of recombinant proteins. Gene 169 53–58 Occurrence Handle8635749 Occurrence Handle10.1016/0378-1119(95)00795-4 Occurrence Handle1:CAS:528:DyaK28XhsVaktrc%3D
E Weiland R Ahl R Stark F Weiland HJ Thiel (1992) ArticleTitleA second envelope glycoprotein mediates neutralization of a pestivirus, hog cholera virus. J Virol 66 3677–3682 Occurrence Handle1583727 Occurrence Handle1:CAS:528:DyaK38Xkt1aqurg%3D
E Weiland R Stark B Haas T Rumenapf G Meyers HJ Thiel (1990) ArticleTitlePestivirus glycoprotein which induces neutralizing antibodies forms part of a disulfide-linked heterodimer. J Virol 64 3563–3569 Occurrence Handle2370675 Occurrence Handle1:CAS:528:DyaK3cXltFensr4%3D
G Wensvoort (1989) ArticleTitleTopographical and functional mapping of epitopes on hog cholera virus with monoclonal antibodies. J Gen Virol 70 2865–2876 Occurrence Handle2479712
JM Windisch R Schneider R Stark E Weiland G Meyers HJ Thiel (1996) ArticleTitleRNase of classical swine fever virus: biochemical characterization and inhibition by virus-neutralizing monoclonal antibodies. J Virol 70 352–358 Occurrence Handle8523547 Occurrence Handle1:CAS:528:DyaK2MXpvFSms7Y%3D
M Yu K Than S Colegate B Shiell WP Michalski S Prowse L-F Wang (2005) ArticleTitlePeptide mimotopes of phomopsins: identification, characterization and application in an immunoassay. Mol Diversity 9 233–240 Occurrence Handle1:CAS:528:DC%2BD2MXhslelsrw%3D
M Yu L-F Wang BJ Shiell CJ Morrissy HA Westbury (1996) ArticleTitleFine mapping of a C-terminal linear epitope highly conserved among the major envelope glycoprotein E2 (gp51 to gp54) of different pestiviruses. Virology 222 289–292 Occurrence Handle8806512 Occurrence Handle10.1006/viro.1996.0423 Occurrence Handle1:CAS:528:DyaK28XkvVCgsLs%3D
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Zhang, F., Yu, M., Weiland, E. et al. Characterization of epitopes for neutralizing monoclonal antibodies to classical swine fever virus E2 and Erns using phage-displayed random peptide library. Arch Virol 151, 37–54 (2006). https://doi.org/10.1007/s00705-005-0623-9
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00705-005-0623-9