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.