The major subunit ClpG of Escherichia coli CS31A fibrillae as an expression vector for different combinations of two TGEV coronavirus epitopes

被引:15
作者
Mechin, MC [1 ]
DerVartanian, M [1 ]
Martin, C [1 ]
机构
[1] INRA,CTR RECH CLERMONT FERRAND THEIX,MICROBIOL LAB,F-63122 ST GENES CHAMPANE,FRANCE
关键词
recombinant DNA; genetic fusion; hybrid protein; tandem insertion; transmissible gastroenteritis virus; peptide presentation; surface exposure;
D O I
10.1016/S0378-1119(96)00348-4
中图分类号
Q3 [遗传学];
学科分类号
071007 ; 090102 ;
摘要
Previously, two B-cell epitopes from the entero-pathogenic transmissible gastroenteritis virus (TGEV), namely the C epitope (TGEV-C) amino acids (aa) 363-371 and the A epitope (TGEV-A) aa 522-531 of the spike S protein (TGEV-S), have been separately expressed on the CS31A fibrillae at the surface of Escherichia coli following insertion into a same region of ClpG. However, the resulting chimeras induced a marginal TGEV-neutralizing antibody (Ab) response in mice. Here, with the view to improving this response, we introduced TGEV-C alone or in different tandem association with TGEV-A (A::C or C::A) in twelve putatively exposed regions of ClpG. Among the 28 resulting engineered proteins only 15, carrying up to 51 extra aa, had not essentially disturbed the correct CS31A fibrillae formation process. Six partially permissive sites accepting only TGEV-C and three highly permissive sites tolerating A::C or C::A tandem peptide, were identified throughout ClpG. Intact bacteria or extracted CS31A hybrid fibrillae expressing TGEV epitopes at any of the permissive sites, were recognized by Ab directed against the foreign parent protein, providing a direct argument for exposure of the corresponding ClpG region at the cell surface and for antigenicity of the epitopes in the polymeric CS31A fibrillae context. The potential of CS31A fibrillae as carriers of the TGEV peptides indicates that there may be three positions (N terminus, aa 202-204 and 202-218) in ClpG which may turn out to be important fusion sites and therefore be relevant for the eventual design of TGEV vaccines. Unexpectedly, TGEV-A, whatever its position in ClpG, mediated the partial proteolytic degradation of the hybrid proteins, suggesting that it functions as a substrate for a cellular protease, and thereby that its suitability as a vaccine antigen candidate is doubtful.
引用
收藏
页码:211 / 218
页数:8
相关论文
共 21 条
[1]   IMPROVED MIMICRY OF A FOOT-AND-MOUTH-DISEASE VIRUS ANTIGENIC SITE BY A VIRAL PEPTIDE DISPLAYED ON BETA-GALACTOSIDASE SURFACE [J].
BENITO, A ;
MATEU, MG ;
VILLAVERDE, A .
BIO-TECHNOLOGY, 1995, 13 (08) :801-804
[2]   CS31A CAPSULE-LIKE ANTIGEN AS AN EXPOSURE VECTOR FOR HETEROLOGOUS ANTIGENIC DETERMINANTS [J].
BOUSQUET, F ;
MARTIN, C ;
GIRARDEAU, JP ;
MECHIN, MC ;
DERVARTANIAN, M ;
LAUDE, H ;
CONTREPOIS, M .
INFECTION AND IMMUNITY, 1994, 62 (06) :2553-2561
[3]   SYNTHESIS OF FUSION PROTEINS WITH MULTIPLE COPIES OF AN ANTIGENIC DETERMINANT OF FOOT-AND-MOUTH-DISEASE VIRUS [J].
BROEKHUIJSEN, MP ;
BLOM, T ;
VANRIJN, J ;
POUWELS, PH ;
KLASEN, EA ;
FASBENDER, MJ ;
ENGERVALK, BE .
GENE, 1986, 49 (02) :189-197
[4]   PERMISSIVE SITES AND TOPOLOGY OF AN OUTER-MEMBRANE PROTEIN WITH A REPORTER EPITOPE [J].
CHARBIT, A ;
RONCO, J ;
MICHEL, V ;
WERTS, C ;
HOFNUNG, M .
JOURNAL OF BACTERIOLOGY, 1991, 173 (01) :262-275
[5]   LOCALIZATION OF ANTIGENIC SITES OF THE E2 GLYCOPROTEIN OF TRANSMISSIBLE GASTROENTERITIS CORONAVIRUS [J].
CORREA, I ;
GEBAUER, F ;
BULLIDO, MJ ;
SUNE, C ;
BAAY, MFD ;
ZWAAGSTRA, KA ;
POSTHUMUS, WPA ;
LENSTRA, JA ;
ENJUANES, L .
JOURNAL OF GENERAL VIROLOGY, 1990, 71 :271-279
[6]   4 MAJOR ANTIGENIC SITES OF THE CORONAVIRUS TRANSMISSIBLE GASTROENTERITIS VIRUS ARE LOCATED ON THE AMINO-TERMINAL HALF OF SPIKE GLYCOPROTEIN-S [J].
DELMAS, B ;
RASSCHAERT, D ;
GODET, M ;
GELFI, J ;
LAUDE, H .
JOURNAL OF GENERAL VIROLOGY, 1990, 71 :1313-1323
[7]   SITE-DIRECTED MUTAGENESIS OF VIRTUALLY ANY PLASMID BY ELIMINATING A UNIQUE SITE [J].
DENG, WP ;
NICKOLOFF, JA .
ANALYTICAL BIOCHEMISTRY, 1992, 200 (01) :81-88
[8]   PERMISSIBLE PEPTIDE INSERTIONS SURROUNDING THE SIGNAL PEPTIDE MATURE PROTEIN JUNCTION OF THE CLPG PREPILIN - CS31A FIMBRIAE OF ESCHERICHIA-COLI AS CARRIERS OF FOREIGN SEQUENCES [J].
DERVARTANIAN, M ;
MECHIN, MC ;
JAFFEUX, B ;
BERTIN, Y ;
FELIX, I ;
GAILLARDMARTINIE, B .
GENE, 1994, 148 (01) :23-32
[9]   RESIDUES INVOLVED IN THE ANTIGENIC SITES OF TRANSMISSIBLE GASTROENTERITIS CORONAVIRUS S-GLYCOPROTEIN [J].
GEBAUER, F ;
POSTHUMUS, WPA ;
CORREA, I ;
SUNE, C ;
SMERDOU, C ;
SANCHEZ, CM ;
LENSTRA, JA ;
MELOEN, RH ;
ENJUANES, L .
VIROLOGY, 1991, 183 (01) :225-238
[10]   CS31A, A NEW K88-RELATED FIMBRIAL ANTIGEN ON BOVINE ENTERO-TOXIGENIC AND SEPTICEMIC ESCHERICHIA-COLI STRAINS [J].
GIRARDEAU, JP ;
DERVARTANIAN, M ;
OLLIER, JL ;
CONTREPOIS, M .
INFECTION AND IMMUNITY, 1988, 56 (08) :2180-2188