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.
National Center for Environmental Health, Centers for Disease Control and Prevention, 4770 Buford Highway, NE, Atlanta, Georgia 30341-3724, USA. aboyer@cdc.gov
Anne E
AE
Boyer
Hercules
H
Moura
Adrian R
AR
Woolfitt
Suzanne R
SR
Kalb
Lisa G
LG
McWilliams
Antonis
A
Pavlopoulos
Jurgen G
JG
Schmidt
David L
DL
Ashley
John R
JR
Barr
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