Ratcheting up protein translocation with anthrax toxin

Abstract

Energy-consuming nanomachines catalyze the directed movement of biopolymers in the cell. They are found both dissolved in the aqueous cytosol as well as embedded in lipid bilayers. Inquiries into the molecular mechanism of nanomachine-catalyzed biopolymer transport have revealed that these machines are equipped with molecular parts, including adjustable clamps, levers, and adaptors, which interact favorably with substrate polypeptides. Biological nanomachines that catalyze protein transport, known as translocases, often require that their substrate proteins unfold before translocation. An unstructured protein chain is likely entropically challenging to bind, push, or pull in a directional manner, especially in a way that produces an unfolding force. A number of ingenious solutions to this problem are now evident in the anthrax toxin system, a model used to study protein translocation. Here we highlight molecular ratchets and current research on anthrax toxin translocation. A picture is emerging of proton-gradientdriven anthrax toxin translocation, and its associated ratchet mechanism likely applies broadly to other systems. We suggest a cyclical thermodynamic order-to-disorder mechanism (akin to a heatengine cycle) is central to underlying protein translocation: peptide substrates nonspecifically bind to molecular clamps, which possess adjustable affinities; polypeptide substrates compress into helical structures; these clamps undergo proton-gated switching; and the substrate subsequently expands regaining its unfolded state conformational entropy upon translocation.

Anthrax toxin-induced rupture of artificial lipid bilayer membranes

“…One model suggests that LF and EF thread through the pore. 19,20 The results shown here suggest that anthrax toxin complexes (i.e., LF or EF bound to the PA 63 channel) rupture membranes. A previous study demonstrated that LF in the complex is enzymatically active.…”

“…Several groups hypothesized that the transport of LF and EF into the cell requires their translocation through the PA 63 pore, 19,20 which has a diameter of ≈1.2-2 nm. [21][22][23] Previous electrophysiological studies demonstrated that full-length LF and EF (≈760 amino acids each) [24][25][26][27] fragments (LF N and EF N , ≈254 amino acids each) [28][29][30][31][32] bind strongly but reversibly to the channel and reduce the pore conductance.…”

Three dimensional structure of the anthrax toxin translocon–lethal factor complex by cryo‐electron microscopy

“…These pores act as translocons, which unfold LF and EF and transport them across the endosomal membrane to the cytosol (see latest review). 1 Prior to structural verification, biochemical and biophysical data indicated that domain 2 (residues 259-487) of the PA prepore rearranges to form a 100 Å -long beta barrel that connects the globular portion of the PA to the target membrane, forming a transmembrane pore. The overall shape of the translocon has been validated using both 2D and ultimately 3D electron microscopy.…”

Anthrax toxin protective antigen integrates poly-γ- d -glutamate and pH signals to sense the optimal environment for channel formation

“…Anthrax toxin (Atx) (13,14)-a virulence factor produced by pathogenic strains of Bacillus anthracis-consists of three nontoxic protein components. The protective antigen (PA) component (83 kDa) binds to cells and forms an oligomeric translocase channel, allowing the other two enzyme components, lethal factor (LF; 90 kDa) and edema factor (EF; 89 kDa), to translocate into the host cell.…”