Localization of the regions on the C-terminal domain of the heavy chain of botulinum A recognized by T lymphocytes and by antibodies after immunization of mice with pentavalent toxoid.
We have mapped the regions recognized by T and/or B cells (Abs) on the C-terminal domain (Hc) of the heavy chain of botulinum neurotoxin serotype A (BoNT/A) after immunization of two inbred mouse strains with pentavalent toxoid (BoNTs A, B, C, D and E). Using a set of synthetic overlapping peptides, encompassing the entire Hc domain (residues 855-1296), we demonstrated that T cells of Balb/c (H-2d) mice, primed with one injection of toxoid, recognized two major regions within residues 897-915 and 939-957. After multiple inoculations with toxoid, T cells of Balb/c expanded their recognition ability and responded very well to challenge with peptide 1261-1279 and moderately to stimulation with peptide 1149-1167. Unlike Balb/c T cells, those of toxoid-primed SJL (H-2s) mice exhibited a more complex profile and responded to challenge with a large number of overlapping peptides. After one toxoid injection, however, three peptides, 897-915, 939-957/953-971 overlap and 1051-1069, were the most potent T cells stimulators. After three toxoid injections, peptides 897-915 and 1051-1069 remained immunodominant while the third region was shifted upstream to 925-943/939-957 overlap. The immunodominant epitope within peptide 897-915 was recognized exclusively by T cells, since no Abs were detected against this region. The Ab binding profiles of the two mouse strains were quite similar, showing only small quantitative differences. Both, Balb/c and SJL anti-toxoid Abs displayed strong binding mainly to peptide 1177-1195, followed by peptides 869-887/883-901 overlap and 1275-1296. In addition, a significant amount of Balb/c anti-toxoid Abs was bound to peptide 1135-1153. Unlike Balb/c Abs, that interacted weakly with peptides 995-1013 and 1051-1069, the anti-toxoid Abs of SJL mice exhibited strong binding toward both peptides. The results showed that, in a given strain, the regions recognized by anti-toxoid Abs and T cells may coincide or may be uniquely B or T cell determinants.
Rosenberg
J S
JS
Department of Biochemistry, Baylor College of Medicine, Houston, TX 77030, USA.
Middlebrook
J L
JL
Atassi
M Z
MZ
D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial; D001428:Bacterial Vaccines / Q000302:isolation & purification; D019274:Botulinum Toxins, Type A / Q000737:chemistry* / Q000235:genetics / Q000276:immunology*; D003014:Clostridium botulinum / Q000276:immunology; D018604:Epitope Mapping; D005260:Female; D007114:Immunization; D008213:Lymphocyte Activation; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology*; D014121:Toxoids / Q000008:administration & dosage
D000907:Antibodies, Bacterial; D001428:Bacterial Vaccines; C035415:Clostridium botulinum toxoid; D010446:Peptide Fragments; D014121:Toxoids; D019274:Botulinum Toxins, Type A