Mapping the lethal factor and edema factor binding sites on oligomeric anthrax protective antigen

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Anthrax toxin assembly at the cell surface is crucial for its toxicity.
• Edema factor (EF) and lethal factor (LF) compete for binding to protective antigen (PA) oligomers.
• Understanding this interaction is key to developing antitoxin strategies.

Purpose of the Study:

• To delineate the binding site for EF and LF on PA(63).
• To elucidate the structural basis of ligand binding and PA oligomerization dependence.
• To provide insights into the assembly mechanism of anthrax toxin.

Main Methods:

• Sequence analysis of PA(63) and related proteins to identify potential binding regions.
• Site-directed mutagenesis of conserved residues within PA domain 1'.
• Oligomerization-deficient PA mutants were used to map the ligand binding site.

Main Results:

• Seven key residues in PA domain 1' were identified as critical for EF and LF binding.
• Mutations in these residues significantly inhibited ligand binding.
• The ligand binding site spans the interface between adjacent PA subunits in dimers and higher-order oligomers.
• The PA(63) heptamer binds a maximum of three ligand molecules.

Conclusions:

• The study elucidates the precise binding site for EF and LF on PA(63).
• Ligand binding is dependent on PA oligomerization, with the site located at subunit interfaces.
• This understanding is vital for designing inhibitors of anthrax toxin assembly and function.
• The findings may apply to the assembly mechanisms of other binary bacterial toxins.