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PLoS One. 2009;4(3):e4924. doi: 10.1371/journal.pone.0004924. Epub 2009 Mar 17.

Epitope characterization and variable region sequence of f1-40, a high-affinity monoclonal antibody to botulinum neurotoxin type a (Hall strain).

Author information

1
United States Department of Agriculture, Agricultural Research Service, Western Regional Research Center, Albany, California, United States of America.

Erratum in

  • PLoS One. 2009;4(10). doi: 10.1371/annotation/6359a6c2-90f8-4f7e-ad86-5bac5d62a7fe.
  • PLoS One. 2009;4(11). doi: 10.1371/annotation/6359a6c2-90f8-4f7e-ad86-5bac5d62a7fe.
  • PLoS One. 2009;4(10). doi: 10.1371/annotation/8d83a0a4-4dd0-4c57-ad94-304130f565e6.
  • PLoS One. 2009;4(10). doi: 10.1371/annotation/47f9bc3f-7c79-453c-85e3-588bc3056951.

Abstract

BACKGROUND:

Botulism, an often fatal neuroparalytic disease, is caused by botulinum neurotoxins (BoNT) which consist of a family of seven serotypes (A-H) produced by the anaerobic bacterium Clostridium botulinum. BoNT, considered the most potent biological toxin known, is a 150 kDa protein consisting of a 100 kDa heavy-chain (Hc) and a 50 kDa light-chain (Lc). F1-40 is a mouse-derived, IgG1 monoclonal antibody that binds the light chain of BoNT serotype A (BoNT/A) and is used in a sensitive immunoassay for toxin detection. We report the fine epitope mapping of F1-40 and the deduced amino acid sequence of the variable regions of the heavy and light chains of the antibody.

METHODS AND FINDINGS:

To characterize the binding epitope of F1-40, three complementary experimental approaches were selected. Firstly, recombinant peptide fragments of BoNT/A light-chain were used in Western blots to identify the epitope domains. Secondly, a peptide phage-display library was used to identify the specific amino acid sequences. Thirdly, the three-dimensional structure of BoNT/A was examined in silico, and the amino acid sequences determined from the phage-display studies were mapped onto the three-dimensional structure in order to visualize the epitope. F1-40 was found to bind a peptide fragment of BoNT/A, designated L1-3, which spans from T125 to L200. The motif QPDRS was identified by phage-display, and was mapped to a region within L1-3. When the three amino acids Q138, P139 and D140 were all mutated to glycine, binding of F1-40 to the recombinant BoNT/A light chain peptide was abolished. Q-138, P-139 and D-140 form a loop on the external surface of BoNT/A, exposed to solvent and accessible to F1-40 binding.

CONCLUSIONS:

The epitope of F1-40 was localized to a single exposed loop (ss4, ss5) on the Lc of BoNT. Furthermore amino acids Q138, P139 and D140 forming the tip of the loop appear critical for binding.

PMID:
19290051
PMCID:
PMC2654115
DOI:
10.1371/journal.pone.0004924
[Indexed for MEDLINE]
Free PMC Article
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