title: Peptide Probes Reveal a Hydrophobic Steric Ratchet in the Anthrax Toxin Protective Antigen Translocase.
abstract: Anthrax toxin is a tripartite virulence factor produced by Bacillus anthracis during infection. Under acidic endosomal pH conditions, the toxin's protective antigen (PA) component forms a transmembrane channel in host cells. The PA channel then translocates its two enzyme components, lethal factor and edema factor, into the host cytosol under the proton motive force. Protein translocation under a proton motive force is catalyzed by a series of nonspecific polypeptide binding sites, called clamps. A 10-residue guest/host peptide model system, KKKKKXXSXX, was used to functionally probe polypeptide-clamp interactions within wild-type PA channels. The guest residues were Thr, Ala, Leu, Phe, Tyr, and Trp. In steady-state translocation experiments, the channel blocked most tightly with peptides that had increasing amounts of nonpolar surface area. Cooperative peptide binding was observed in the Trp-containing peptide sequence but not the other tested sequences. Trp substitutions into a flexible, uncharged linker between the lethal factor amino-terminal domain and diphtheria toxin A chain expedited translocation. Therefore, peptide-clamp sites in translocase channels can sense large steric features (like tryptophan) in peptides, and while these steric interactions may make a peptide translocate poorly, in the context of folded domains, they can make the protein translocate more rapidly presumably via a hydrophobic steric ratchet mechanism.
authors.affiliation: Molecular Toxicology Graduate Program, University of California, Berkeley, CA 94720, USA.
authors.forename: Jennifer M
authors.initials: JM
authors.lastname: Colby
authors.affiliation: Department of Chemistry, University of California, Berkeley, CA 94720, USA; Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA; Department of Microbial Pathogenesis, School of Dentistry, University of Maryland, 650 West Baltimore St., Baltimore, MD 21201, USA. Electronic address: bkrantz@umaryland.edu.
authors.forename: Bryan A
authors.initials: BA
authors.lastname: Krantz
mesh_terms: D022122:Anthrax Vaccines / Q000737:chemistry / Q000378:metabolism; D000942:Antigens, Bacterial / Q000737:chemistry* / Q000378:metabolism*; D001408:Bacillus anthracis / Q000201:enzymology*; D001427:Bacterial Toxins / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism*; D004594:Electrophysiology; D057927:Hydrophobic and Hydrophilic Interactions; D008958:Models, Molecular; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism*; D021381:Protein Transport
chemical_list: D022122:Anthrax Vaccines; D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D010446:Peptide Fragments; C030325:anthrax toxin
keywords: Bacillus anthracis; anthrax toxin; electrophysiology; protective antigen; ratchet