Biologically active novel conformational state of botulinum, the most poisonous poison.
Botulinum neurotoxins (serotypes A-G), the most toxic substances known to humankind, cause flaccid muscle paralysis by blocking acetylcholine release at nerve-muscle junctions through a very specific and exclusive endopeptidase activity against SNARE proteins of presynaptic exocytosis machinery. We have examined polypeptide folding of the endopeptidase moiety of botulinum neurotoxin/A (the light chain) under conditions of its optimal enzymatic activity and have found that one of its stable conformational states is a molten-globule, which retains over 60% of its optimal enzyme activity. More importantly, we have discovered that the light chain acquires a novel pre-imminent molten-globule enzyme conformation at the physiologically relevant temperature, 37 degrees C. The pre-imminent molten-globule enzyme form also exhibited the maximum endopeptidase activity against its intracellular substrate, SNAP-25 (synaptosomal associated protein of 25 kDa). These findings will not only open new avenues to design effective diagnostics and antidotes against botulism but also provide new information on enzymatically active molten-globule or molten-globule like structures.
Kukreja
Roshan
R
Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, Dartmouth, Massachusetts 02747, USA.
Singh
BalRam
B
D019274:Botulinum Toxins, Type A / Q000737:chemistry* / Q000378:metabolism / Q000633:toxicity*; D002942:Circular Dichroism; D010450:Endopeptidases / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D008958:Models, Molecular; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity; D013056:Spectrophotometry, Ultraviolet; D013379:Substrate Specificity; D050825:Synaptosomal-Associated Protein 25 / Q000737:chemistry / Q000378:metabolism; D013696:Temperature
D011994:Recombinant Proteins; C494208:SNAP25 protein, human; D050825:Synaptosomal-Associated Protein 25; D010450:Endopeptidases; D019274:Botulinum Toxins, Type A