Anti-B. anthracis Lethal Factor Monoclonal antibody, Clone CAL0106

Inhalational anthrax is caused by B. anthracis, a virulent spore-forming bacterium which secretes anthrax toxins consisting of protective antigen (PA), lethal factor (IF) and edema factor (EF). LF is a Zn-dependent metalloprotease and is the main determinant in the pathogenesis of anthrax. Here we report the identification of a lead small-molecule inhibitor of anthrax lethal factor by screening an available synthetic small-molecule inhibitor library using fluorescence-based high-throughput screening (HIS) approach. Seven small molecules were found to have inhibitory effect against IF activity, among which SM157 had the highest inhibitory activity. All theses small molecule inhibitors inhibited IS in a noncompetitive inhibition mode. SM157 and SM167 are from the same family, both having an identical group complex, which is predicted to insert into S1' pocket of LE. More potent small-molecule inhibitors could be developed by modifying SM157 based on this identical group complex. [BMB reports 2011; 44(12): 811-815]

Anthrax lethal factor (LF) is a Zn2+-metalloprotease that cleaves and inactivates mitogen-activated protein kinase kinases (MEKs). We have used site-directed mutagenesis to identify a cluster of residues in domain II of LF that lie outside the active site and are required for cellular proteolytic activity toward MEKs. Alanine substituted for Leu(293), Lys(294), Leu(514), Asn(516), or Arg(491) caused a 10-50-fold reduction in LF toxicity. Further, whereas pairwise substitution of alanine for Leu(514) and either Leu(293), Lys(294), or Arg(491) completely abrogated LF toxicity, pairwise mutation of Leu(514) and Asn(516) resulted in toxicity comparable with N516A alone. The introduction of these mutations reduced LF-mediated cleavage of MEK2 in cell-based assays but altered neither the ability of LF to bind protective antigen nor its ability to translocate across a membrane. Interestingly, direct in vitro measurement of LF activity indicated that decreased toxicity was not always accompanied by reduced proteolytic activity. However, mutations in this region significantly reduced the ability of LF to competitively inhibit B-Raf phosphorylation of MEK. These results provide evidence that elements of domain II are involved in the association of LF into productive complex with MEKs.