Immunological characterization of the subunits of type A botulinum neurotoxin and different components of its associated proteins.
Botulinum neurotoxins (BoNTs) constitute a family of seven structurally similar but antigenically distinct proteins produced by different strains of Clostridium botulinum. Type A botulinum neurotoxin (BoNT/A) is produced along with 6 neurotoxin associated proteins (NAPs) including hemagglutinin (Hn-33) through polycistronic expression of a clustered group of genes to form a complex (BoNT/AC). The presence of NAPs enhances the oral toxicity of the neurotoxin significantly. Hn-33 makes up the largest fraction of NAPs in BoNT/AC and strongly protects BoNT/A against proteases of the GI tract. BoNT in its complex form is also used in therapeutic and cosmetic applications to treat several neuromuscular disorders. In this study immunological reactivity of BoNT/A in its purified and complex forms, neurotoxin associated proteins, and Hn-33 have been examined using enzyme-linked immunosorbent assay (ELISA). Antibodies raised against the whole complex reacted 60 times better with the complex and 35 times better with Hn-33 and NAPs compared to the purified neurotoxin suggesting stronger immunogenicity of NAPs over that of purified neurotoxin and a higher potential of BoNT/AC and its associated proteins to induce host immune response. This observation also suggests that Hn-33 and other NAPs could potentially be employed as adjuvants for development of vaccines against botulism and could be a good surrogate for botulinum diagnostics. ELISA binding curves of BoNT/AC and BoNT/A with antibodies raised against BoNT/A indicate that BoNT/A in its purified and complex forms induces equal immunogenic response and a 2.5-fold higher immunogenic response compared to BoNT/A light and heavy chains. We have also discovered a new protein, an intimin analog, present within the complex preparation of BoNT/A which shows dramatically high immunoreactivity.
Kukreja
Roshan
R
Botulinum Research Center, University of Massachusetts Dartmouth, 285 Old Westport Road, North Dartmouth, MA 02747, USA.
Chang
Tzuu-Wang
TW
Cai
Shuowei
S
Lindo
Paul
P
Riding
Stephen
S
Zhou
Yu
Y
Ravichandran
Easwaran
E
Singh
Bal Ram
BR
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000276:immunology*; D019274:Botulinum Toxins, Type A / Q000737:chemistry / Q000276:immunology*; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D021122:Protein Subunits; D011817:Rabbits
D001426:Bacterial Proteins; D021122:Protein Subunits; D019274:Botulinum Toxins, Type A