title: Beltless translocation domain of botulinum neurotoxin A embodies a minimum ion-conductive channel.
abstract: Botulinum neurotoxin, the causative agent of the paralytic disease botulism, is an endopeptidase composed of a catalytic domain (or light chain (LC)) and a heavy chain (HC) encompassing the translocation domain (TD) and receptor-binding domain. Upon receptor-mediated endocytosis, the LC and TD are proposed to undergo conformational changes in the acidic endocytic environment resulting in the formation of an LC protein-conducting TD channel. The mechanism of channel formation and the conformational changes in the toxin upon acidification are important but less well understood aspects of botulinum neurotoxin intoxication. Here, we have identified a minimum channel-forming truncation of the TD, the "beltless" TD, that forms transmembrane channels with ion conduction properties similar to those of the full-length TD. At variance with the holotoxin and the HC, channel formation for both the TD and the beltless TD occurs independent of a transmembrane pH gradient. Furthermore, acidification in solution induces moderate secondary structure changes. The subtle nature of the conformational changes evoked by acidification on the TD suggests that, in the context of the holotoxin, larger structural rearrangements and LC unfolding occur preceding or concurrent to channel formation. This notion is consistent with the hypothesis that although each domain of the holotoxin functions individually, each domain serves as a chaperone for the others.
authors.affiliation: Section of Neurobiology, Division of Biological Sciences, University of California at San Diego, La Jolla, California 92093-0366, USA.
authors.forename: Audrey
authors.initials: A
authors.lastname: Fischer
authors.forename: Shilpa
authors.initials: S
authors.lastname: Sambashivan
authors.forename: Axel T
authors.initials: AT
authors.lastname: Brunger
authors.forename: Mauricio
authors.initials: M
authors.lastname: Montal
mesh_terms: D019274:Botulinum Toxins, Type A / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001906:Botulism / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D006801:Humans; D007473:Ion Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010449:Peptide Mapping / Q000379:methods; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018892:Proton-Motive Force
chemical_list: D007473:Ion Channels; D018832:Molecular Chaperones; D019274:Botulinum Toxins, Type A