Cryo-Em Structure Of The Anthrax Toxin Protective Antigen Channel Bound To Lethal Factor

Anthrax toxin consists of three individual protein factors: the translocase channel forming protective antigen (PA), and the enzymatic factors lethal factor (LF) and edema factor (EF). Anthrax toxin is an excellent model system for studying the biophysical mechanisms of protein translocation. Binding of LF/EF to the PA pre-channel serves to stabilize their unfolding before translocation. There is limited high-resolution structural data of substrate binding, specifically of the amino-terminal domain of LF (LFn), to the octameric PA pre-channel. We were able to determine the high-resolution structure of the heptameric PA in its pore conformation bound to LF at 4.6-Å. Using lipid nanodisc, we were able to obtain random orientation of our PA toxin complex on a cryo-EM grid and therefore perform single-particle analysis and structure determination. The PA7LF3 channel complex was inserted into lipid nanodiscs. The structure contains the PA heptamer bound to a single LF. Under these conditions, the C-Terminal domains of LF appears to be disordered, resulting in no electron density for this region of the protein. There appears to be no difference in the structure of the visible LFn domain when bound to PA in the pore state vs. the pre-channel complex, with the first α-helix and β-sheet in LF unfolded and docked within the α-clamp site. In addition, there appears to be a density present within the phenylalanine clamp (Φ-clamp) site. Further studies are currently being performed in an effort to identify this density in an attempt to provide structural insight into the mechanisms by which proteins translocate through this PA channel.