title: Role of the disulfide cleavage induced molten globule state of type a botulinum neurotoxin in its endopeptidase activity.
abstract: Botulinum neurotoxins are produced by anaerobic Clostridium botulinum in an inactive form. The endopeptidase activity of type A botulinum neurotoxin (BoNT/A) is triggered by reduction of its disulfide bond between its heavy chain and light chain. By using circular dichroism spectroscopy, we show that, upon reduction of BoNT/A and under physiological temperature (37 degrees C), the BoNT/A loses most of its native tertiary structure, while retaining most of its secondary structure. This type of structure is characterized as a molten globule type conformation, which was further confirmed for BoNT/A by the characteristic binding of 1-anilinonaphthalene-8-sulfonic acid. Under nonreducing conditions where the interchain disulfide bond is intact, the enzymatically inactive BoNT/A did not show a molten globule type of structure. A temperature profile of the structure and enzyme activity of BoNT/A revealed that, under reducing conditions, there was a strong correlation in the existence of the molten globule structure and optimum endopeptidase activity at about 37 degrees C.
authors.affiliation: Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, Massachusetts 02747, USA.
authors.forename: S
authors.initials: S
authors.lastname: Cai
authors.forename: B R
authors.initials: BR
authors.lastname: Singh
mesh_terms: D000817:Anilino Naphthalenesulfonates; D019274:Botulinum Toxins, Type A / Q000737:chemistry* / Q000378:metabolism*; D002942:Circular Dichroism; D004220:Disulfides / Q000737:chemistry; D010450:Endopeptidases / Q000737:chemistry* / Q000378:metabolism*; D005456:Fluorescent Dyes; D008958:Models, Molecular; D010084:Oxidation-Reduction; D017434:Protein Structure, Tertiary
chemical_list: D000817:Anilino Naphthalenesulfonates; D004220:Disulfides; D005456:Fluorescent Dyes; C027132:1-anilino-8-naphthalenesulfonate; D010450:Endopeptidases; D019274:Botulinum Toxins, Type A