Improved Detection of Botulinum Neurotoxin Serotype A by Endopep-MS through Peptide Substrate Modification

Details

• Alternative Title:
  Anal Biochem
• Personal Author:
  Wang, Dongxia ; Baudys, Jakub ; Ye, Yiming ; Rees, Jon C. ; Barr, John R. ; Pirkle, James L. ; Kalb, Suzanne R.
• Description:
  Botulinum neurotoxins (BoNTs) are a family of seven toxin serotypes that are the most toxic substances known to humans. Intoxication with BoNT causes flaccid paralysis and can lead to death if untreated with serotype-specific antibodies. Supportive care, including ventilation, may be necessary. Rapid and sensitive detection of BoNT is necessary for timely clinical confirmation of clinical botulism. Previously, our laboratory developed a fast and sensitive mass spectrometry (MS) method termed the Endopep-MS assay. The BoNT serotypes are rapidly detected and differentiated by extracting the toxin with serotype-specific antibodies and detecting the unique and serotype-specific cleavage products of peptide substrates that mimic the sequence of the BoNT native targets. To further improve the sensitivity of the Endopep-MS assay, we report here the optimization of the substrate peptide for the detection of BoNT/A. Modifications on the terminal groups of the original peptide substrate with acetylation and amidation significantly improved the detection of BoNT/A cleavage products. The replacement of some internal amino acid residues with single or multiple substitutions led to further improvement. An optimized peptide increased assay sensitivity 5-fold with toxin spiked into buffer solution or different biological matrices.
• Subjects:
  Acetylation Amino Acid Sequence Botulinum Toxins, Type A Botulism Endopeptidases Immunoglobulin G Molecular Sequence Data Peptides Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization Substrate Specificity
• Source:
  Anal Biochem. 432(2):115-123.
• Pubmed ID:
  23017875
• Pubmed Central ID:
  PMC4618377
• Document Type:
  Journal Article
• Funding:
  CC999999/Intramural CDC HHS/United States
• Volume:
  432
• Issue:
  2
• Collection(s):
  CDC Public Access
• Main Document Checksum:
  urn:sha256:a2bc1346af54010464e2e2a485b2d0441bf5f2d6762b6b46d4c23ef7016a3550
• Download URL:
  https://stacks.cdc.gov/view/cdc/35312/cdc_35312_DS1.pdf
• File Type:
  [PDF - 1.19 MB ]