Document Detail

Identification of novel small molecules that bind to two different sites on the surface of tetanus toxin C fragment.
MedLine Citation:
PMID:  12387617     Owner:  NLM     Status:  MEDLINE    
A combination of computational methods, electrospray ionization mass spectroscopy (ESI-MS), and NMR spectroscopy has been used to identify novel small molecules that bind to two adjacent sites on the surface of the C fragment of tetanus toxin (TetC). One of these sites, Site-1, binds gangliosides present on the surface of motor neurons, while Site-2 is a highly conserved deep cleft in the structures of the tetanus (TeNT) and botulinum (BoNT) neurotoxins. ESI-MS was used to experimentally determine which of the top 11 computationally predicted Site-2 candidates bind to TetC. Each of the six molecules that tested positive was further screened, individually and as mixtures, for binding to TetC in aqueous solutions by NMR. A trNOESY competition assay was developed that used doxorubicin as a marker for Site-1 to provide insight into whether the predicted Site-2 ligands bound to a different site. Of the six predicted Site-2 ligands tested, only four were observed to bind. Naphthofluorescein-di-beta-galactopyranoside was insoluble under conditions compatible with TetC. Sarcosine-Arg-Gly-Asp-Ser-Pro did not appear to bind, but its binding affinity may have been outside the range detectable by the trNOESY experiment. Of the remaining four, three [3-(N-maleimidopropionyl)biocytin, lavendustin A, and Try-Glu-Try] bind in the same site, presumably the predicted Site-2. The fourth ligand, Ser-Gln-Asn-Tyr-Pro-Ile-Val, binds in a third site that differs from Site-1 or predicted Site-2. The results provide a rational, cost- and time-effective strategy for the selection of an optimal set of Site-1 binders and predicted Site-2 binders for use in synthesizing novel bidendate antidotes or detection reagents for clostridial neurotoxins, such as TeNT and BoNT.
Monique Cosman; Felice C Lightstone; V V Krishnan; Loreen Zeller; Maria C Prieto; Diana C Roe; Rod Balhorn
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.    
Journal Detail:
Title:  Chemical research in toxicology     Volume:  15     ISSN:  0893-228X     ISO Abbreviation:  Chem. Res. Toxicol.     Publication Date:  2002 Oct 
Date Detail:
Created Date:  2002-10-21     Completed Date:  2003-04-17     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  8807448     Medline TA:  Chem Res Toxicol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  1218-28     Citation Subset:  IM    
Biology and Biotechnology Research Program, Lawrence Livermore National Laboratory, Livermore, California 94551-0808, USA.
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MeSH Terms
Binding Sites
Clostridium / pathogenicity
Drug Design
Magnetic Resonance Spectroscopy
Molecular Structure
Motor Neurons
Peptide Fragments / analysis,  chemistry*
Spectrometry, Mass, Electrospray Ionization
Tetanus Toxin / analysis,  chemistry*
Reg. No./Substance:
0/Antidotes; 0/Ligands; 0/Peptide Fragments; 0/Tetanus Toxin; 0/tetanus toxin fragment C

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