Purified Bacillus anthracis lethal toxin complex formed in vitro and during infection exhibits functional and biological activity.

Abstract

Anthrax protective antigen (PA, 83 kDa), a pore-forming protein, upon protease activation to 63 kDa (PA(63)), translocates lethal factor (LF) and edema factor (EF) from endosomes into the cytosol of the cell. The relatively small size of the heptameric PA(63) pore (approximately 12 angstroms) raises questions as to how large molecules such as LF and EF can move through the pore. In addition, the reported high binding affinity between PA and EF/LF suggests that EF/LF may not dissociate but remain complexed with activated PA(63). In this study, we found that purified (PA(63))(7)-LF complex exhibited biological and functional activities similar to the free LF. Purified LF complexed with PA(63) heptamer was able to cleave both a synthetic peptide substrate and endogenous mitogen-activated protein kinase kinase substrates and kill susceptible macrophage cells. Electrophysiological studies of the complex showed strong rectification of the ionic current at positive voltages, an effect similar to that observed if LF is added to the channels formed by heptameric PA(63) pore. Complexes of (PA(63))(7)-LF found in the plasma of infected animals showed functional activity. Identifying active complex in the blood of infected animals has important implications for therapeutic design, especially those directed against PA and LF. Our studies suggest that the individual toxin components and the complex must be considered as critical targets for anthrax therapeutics.

• Authors: Rekha G Panchal, Kelly M Halverson, Wilson Ribot, Douglas Lane, Tara Kenny, Teresa G Abshire, John W Ezzell, Timothy A Hoover, Bradford Powell, Stephen Little, John J Kasianowicz, Sina Bavari
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 280 Issue 11 Pg. 10834-9 (Mar 18 2005) ISSN: 0021-9258 [Print] United States
• PMID: 15644338 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, Non-P.H.S.)

Chemical References

• Antigens, Bacterial
• Bacterial Toxins
• Lipid Bilayers
• Peptides
• anthrax toxin
• Mitogen-Activated Protein Kinase Kinases

Topics

• Animals
• Anthrax (metabolism)
• Antigens, Bacterial (chemistry, metabolism)
• Bacillus anthracis (metabolism)
• Bacterial Toxins (chemistry, metabolism)
• Blotting, Western
• Chromatography, High Pressure Liquid
• Dose-Response Relationship, Drug
• Electrophysiology
• Guinea Pigs
• In Vitro Techniques
• Kinetics
• Lipid Bilayers
• Mitogen-Activated Protein Kinase Kinases (chemistry)
• Peptides (chemistry)
• Protein Binding
• Rabbits
• Substrate Specificity
• Temperature
• Time Factors