The three separate proteins that make up anthrax toxin-protective antigen (PA), edema factor (EF) and lethal factor (LF) act in binary combinations to produce two distinct reactions in experimental animals: edema (PA+EF) and death (PA+LF). PA is believed to interact with a membrane receptor and, after proteolytic processing, to mediate endocytosis and subsequent translocation of EF or LF into the cytosol. Residues W346, M350, and L352 in loop 3 of domain 2 have been implicated to induce a conformational change when the pH is lowered from 7.4 to 6.5. Modification of the residues Trp (346), Met (350), and Leu (352) to alanine individually and all the three residues together to alanine residues resulted in the loss of cytotoxic activity in combination with LF. The mutant proteins were able to bind to the cell surface receptor, become cleaved by trypsin, bind LF, and oligomerize. These residues might play an important role in the membrane insertion of PA and/or translocation of LF/EF into the cytosol.