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Mass Spectrometric Detection of Bacterial Protein Toxins and Their Enzymatic Activity.

Kalb SR, Boyer AE, Barr JR - Toxins (Basel) (2015)

Bottom Line: One such example is botulinum neurotoxin (BoNT), a potent neurotoxin produced by C. botulinum.There are seven known serotypes of BoNT, A-G, and many of the serotypes can be further differentiated into toxin variants, which are up to 99.9% identical in some cases.Mass spectrometric proteomic techniques have been established to differentiate the serotype or toxin variant of BoNT produced by varied strains of C. botulinum.

View Article: PubMed Central - PubMed

Affiliation: Centers for Disease Control and Prevention, 4770 Buford Hwy NE, Atlanta, GA 30341, USA. skalb@cdc.gov.

ABSTRACT
Mass spectrometry has recently become a powerful technique for bacterial identification. Mass spectrometry approaches generally rely upon introduction of the bacteria into a matrix-assisted laser-desorption time-of-flight (MALDI-TOF) mass spectrometer with mass spectrometric recognition of proteins specific to that organism that form a reliable fingerprint. With some bacteria, such as Bacillus anthracis and Clostridium botulinum, the health threat posed by these organisms is not the organism itself, but rather the protein toxins produced by the organisms. One such example is botulinum neurotoxin (BoNT), a potent neurotoxin produced by C. botulinum. There are seven known serotypes of BoNT, A-G, and many of the serotypes can be further differentiated into toxin variants, which are up to 99.9% identical in some cases. Mass spectrometric proteomic techniques have been established to differentiate the serotype or toxin variant of BoNT produced by varied strains of C. botulinum. Detection of potent biological toxins requires high analytical sensitivity and mass spectrometry based methods have been developed to determine the enzymatic activity of BoNT and the anthrax lethal toxins produced by B. anthracis. This enzymatic activity, unique for each toxin, is assessed with detection of the toxin-induced cleavage of strategically designed peptide substrates by MALDI-TOF mass spectrometry offering unparalleled specificity. Furthermore, activity assays allow for the assessment of the biological activity of a toxin and its potential health risk. Such methods have become important diagnostics for botulism and anthrax. Here, we review mass spectrometry based methods for the enzymatic activity of BoNT and the anthrax lethal factor toxin.

No MeSH data available.


Related in: MedlinePlus

MALDI-TOF mass spectra indicating the presence of (A) BoNT/A; (B) /B; (C) /E; and (D) /F. Peptide cleavage products indicating the presence of the neurotoxin are marked with asterisks.
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toxins-07-03497-f004: MALDI-TOF mass spectra indicating the presence of (A) BoNT/A; (B) /B; (C) /E; and (D) /F. Peptide cleavage products indicating the presence of the neurotoxin are marked with asterisks.

Mentions: Beyond identification of the presence of BoNT through proteomics methods, mass spectrometry can be used to determine the enzymatic activity of the toxin to offer an accurate measurement of the health threat of BoNT. Our laboratory first reported this method in 2005 with incubation of BoNT with a peptide substrate similar to the toxin’s natural target, SNAP-25 or synaptobrevin-2 [32,33]. Although BoNT/A, /C, and /E cleave SNAP-25 and /B, /D, /F and /G cleave synaptobrevin-2, each serotype of BoNT cleaves in a different location, as seen in Figure 1. Thus, peptide substrates incorporating the different cleavage locations of BoNTs will be cleaved in a serotype-specific fashion. The peptide mixture is analyzed by MALDI-TOF mass spectrometry, which accurately reports the mass of peptides, both cleaved and intact. Detection of peptide cleavage products corresponding to a particular cleavage site indicates the presence of a specific BoNT serotype. Conversely, if the peptide substrate either remains intact or is cleaved in a location other than the toxin-specific site, then that BoNT serotype is absent. The amino acid sequences of the peptide substrates are listed in Table 1, and Figure 4 depicts the cleavage of peptide substrates indicating the presence of BoNT/A, /B, /E, and /F.


Mass Spectrometric Detection of Bacterial Protein Toxins and Their Enzymatic Activity.

Kalb SR, Boyer AE, Barr JR - Toxins (Basel) (2015)

MALDI-TOF mass spectra indicating the presence of (A) BoNT/A; (B) /B; (C) /E; and (D) /F. Peptide cleavage products indicating the presence of the neurotoxin are marked with asterisks.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591662&req=5

toxins-07-03497-f004: MALDI-TOF mass spectra indicating the presence of (A) BoNT/A; (B) /B; (C) /E; and (D) /F. Peptide cleavage products indicating the presence of the neurotoxin are marked with asterisks.
Mentions: Beyond identification of the presence of BoNT through proteomics methods, mass spectrometry can be used to determine the enzymatic activity of the toxin to offer an accurate measurement of the health threat of BoNT. Our laboratory first reported this method in 2005 with incubation of BoNT with a peptide substrate similar to the toxin’s natural target, SNAP-25 or synaptobrevin-2 [32,33]. Although BoNT/A, /C, and /E cleave SNAP-25 and /B, /D, /F and /G cleave synaptobrevin-2, each serotype of BoNT cleaves in a different location, as seen in Figure 1. Thus, peptide substrates incorporating the different cleavage locations of BoNTs will be cleaved in a serotype-specific fashion. The peptide mixture is analyzed by MALDI-TOF mass spectrometry, which accurately reports the mass of peptides, both cleaved and intact. Detection of peptide cleavage products corresponding to a particular cleavage site indicates the presence of a specific BoNT serotype. Conversely, if the peptide substrate either remains intact or is cleaved in a location other than the toxin-specific site, then that BoNT serotype is absent. The amino acid sequences of the peptide substrates are listed in Table 1, and Figure 4 depicts the cleavage of peptide substrates indicating the presence of BoNT/A, /B, /E, and /F.

Bottom Line: One such example is botulinum neurotoxin (BoNT), a potent neurotoxin produced by C. botulinum.There are seven known serotypes of BoNT, A-G, and many of the serotypes can be further differentiated into toxin variants, which are up to 99.9% identical in some cases.Mass spectrometric proteomic techniques have been established to differentiate the serotype or toxin variant of BoNT produced by varied strains of C. botulinum.

View Article: PubMed Central - PubMed

Affiliation: Centers for Disease Control and Prevention, 4770 Buford Hwy NE, Atlanta, GA 30341, USA. skalb@cdc.gov.

ABSTRACT
Mass spectrometry has recently become a powerful technique for bacterial identification. Mass spectrometry approaches generally rely upon introduction of the bacteria into a matrix-assisted laser-desorption time-of-flight (MALDI-TOF) mass spectrometer with mass spectrometric recognition of proteins specific to that organism that form a reliable fingerprint. With some bacteria, such as Bacillus anthracis and Clostridium botulinum, the health threat posed by these organisms is not the organism itself, but rather the protein toxins produced by the organisms. One such example is botulinum neurotoxin (BoNT), a potent neurotoxin produced by C. botulinum. There are seven known serotypes of BoNT, A-G, and many of the serotypes can be further differentiated into toxin variants, which are up to 99.9% identical in some cases. Mass spectrometric proteomic techniques have been established to differentiate the serotype or toxin variant of BoNT produced by varied strains of C. botulinum. Detection of potent biological toxins requires high analytical sensitivity and mass spectrometry based methods have been developed to determine the enzymatic activity of BoNT and the anthrax lethal toxins produced by B. anthracis. This enzymatic activity, unique for each toxin, is assessed with detection of the toxin-induced cleavage of strategically designed peptide substrates by MALDI-TOF mass spectrometry offering unparalleled specificity. Furthermore, activity assays allow for the assessment of the biological activity of a toxin and its potential health risk. Such methods have become important diagnostics for botulism and anthrax. Here, we review mass spectrometry based methods for the enzymatic activity of BoNT and the anthrax lethal factor toxin.

No MeSH data available.


Related in: MedlinePlus