Details:

Alternative Title:
Anal Chem
Personal Author:
Wang, Dongxia ; Baudys, Jakub ; Rees, Jon ; Marshall, Kristin M. ; Kalb, Suzanne R. ; ... More +
Wang, Dongxia ; Baudys, Jakub ; Rees, Jon ; Marshall, Kristin M. ; Kalb, Suzanne R. ; Parks, Bryan A. ; Nowaczyk, Louis ; Pirkle, James L. ; Barr, John R. Less -
Description:
Botulinum neurotoxin (BoNT) is one of the most toxic substances known. BoNT is classified into seven distinct serotypes labeled A-G. Among individual serotypes, researchers have identified subtypes based on amino acid variability within a serotype and toxin variants with minor amino acid sequence differences within a subtype. BoNT subtype identification is valuable for tracing and tracking bacterial pathogens. A proteomics approach is useful for BoNT subtyping since botulism is caused by botulinum neurotoxin and does not require the presence of the bacteria or its DNA. Enzymatic digestion and peptide identification using tandem mass spectrometry determines toxin protein sequences. However, with the conventional one-step digestion method, producing sufficient numbers of detectable peptides to cover the entire protein sequence is difficult, and incomplete sequence coverage results in uncertainty in distinguishing BoNT subtypes and toxin variants because of high sequence similarity. We report here a method of multiple enzymes and sequential in-gel digestion (MESID) to characterize the BoNT protein sequence. Complementary peptide detection from toxin digestions has yielded near-complete sequence coverage for all seven BoNT serotypes. Application of the method to a BoNT-contaminated carrot juice sample resulted in the identification of 98.4% protein sequence which led to a confident determination of the toxin subtype.
Subject:
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Amino Acid Sequence
Article
Botulinum Toxins
Chromatography, Liquid
Clostridium Botulinum
Endopeptidases
Gels
Guanidine
Molecular Sequence Data
Molecular Typing
Neurotoxins
Proteomics
Sequence Alignment
Tandem Mass Spectrometry

Source:
Anal Chem. 84(11):4652-4658
Pubmed ID:
22577857
Pubmed Central ID:
PMC4582766
Document Type:
Journal Article
Funding:
CC999999/Intramural CDC HHS/United States
Collection(s):
CDC Public Access
Main Document Checksum:
[+]
urn:sha256:3444ef6c6f8a0e0d9a79848e001b705f7997c5dc3de8d9ae58deec3802b24a0c
File Type:

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