Expression and purification of functional human anthrax toxin receptor (ATR/TEM8) binding domain from Escherichia coli.

Anthrax is caused by the gram-positive, spore-forming bacterium, Bacillus anthracis. Anthrax receptors play a crucial role in the pathogenesis of the anthrax disease. Anthrax toxin receptor ATR/TEM8 VWA domain is responsible for the binding of protective antigen (PA) of B. anthracis, and thus an attractive target for structure-based drug therapies. However, the production of soluble and functional ATR/TEM8 VWA domain currently requires the use of mammalian expression systems. In this work, we expressed the ATR/TEM8 VWA domain as a fusion protein in Escherichia coli. Recombinant ATR/TEM8 VWA domain has been purified to homogeneity, and its identity has been verified by both N-terminal protein microsequencing and mass spectrometry. The purified ATR/TEM8 VWA domain exhibits very high affinity to PA based on BIAcore assay. Moreover, like the domain expressed in mammalian system, the bacterially expressed ATR/TEM8 VWA domain can block cytotoxicity induced by anthrax toxins, suggesting that the bacterially expressed ATR/TEM8 VWA domain is properly folded and fully functional.