From the mouse to the mass spectrometer: detection and differentiation of the endoproteinase activities of botulinum neurotoxins A-G by mass spectrometry.
We have developed an assay (Endopep-MS) that detects the specific endoproteinase activities of all seven BoNT types by mass spectrometry (MS). Each BoNT type cleaves a unique site on proteins involved in neuronal transmission. Target peptide substrates based on these proteins identify a BoNT type by its enzymatic action on the substrate and the production of two peptide products, which are then detected by matrix-assisted laser desorption/ionization time-of-flight MS or liquid chromatography electrospray ionization MS/MS. We showed the ability to detect all seven toxin types in a multiplexed assay format. The detection limits achieved range from 0.039 to 0.625 mouse LD(50)/mL for toxin types A, B, E, and F in a buffer system. The Endopep-MS assay is the first to differentiate all seven BoNT types, is sensitive, specific, and has the potential to quantify toxin activity.
Boyer
Anne E
AE
National Center for Environmental Health, Centers for Disease Control and Prevention, 4770 Buford Highway, NE, Atlanta, Georgia 30341-3724, USA. aboyer@cdc.gov
Moura
Hercules
H
Woolfitt
Adrian R
AR
Kalb
Suzanne R
SR
McWilliams
Lisa G
LG
Pavlopoulos
Antonis
A
Schmidt
Jurgen G
JG
Ashley
David L
DL
Barr
John R
JR
D000595:Amino Acid Sequence; D000818:Animals; D001905:Botulinum Toxins / Q000737:chemistry / Q000378:metabolism*; D010450:Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D010450:Endopeptidases; D001905:Botulinum Toxins