Microtech2010 2010

Anthrax Biosensor based on Protective Antigen Ion Channel

A. Michelman-Ribeiro, V. Silin, J. Robertson, S. Kreuger, J. Reiner, J. Kasianowicz
NIST, US

Keywords: anthrax, ion channel, biosensor

Abstract:

Bioterrorism involving anthrax is a reality in the U.S. An important capability is to detect and identify the presence of Bacillus anthracis and anthrax Lethal Toxins. B. anthracis produces three toxins, which are involved in late-stage anthrax infection: Protective Antigen (PA), Lethal Factor (LF), and Edema Factor (EF). Protective Antigen binds to receptors on the cell membrane, is cleaved into PA63, oligomerizes into a heptameric prepore, which then binds LF or EF, and this complex is endocytosed. PA63 forms a membrane channel in the endocytic vesicle, and catalyzes the entry of LF and EF into the cytosol, where they disrupt enzymatic pathways and lead to cell death. We previously demonstrated that LF and EF bind to and block the PA63 channel. We have developed a tethered lipid bilayer system that allows the functional reconstitution of PA63 channels, as judged by surface plasmon resonance and electrochemical impedance spectroscopy. We have used this system, in conjunction with electrophysiology on unsupported bilayers, to optimize both PA63 channel formation and its use as a sensor for LF and EF. The device should also prove useful for the rapid screening of therapeutic agents against anthrax Lethal Toxin.
 
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