Fig. 5. Disturbance of the interaction between 2β3−2β4 loop of PA and Glu-122 pocket of CMG2 lowers the pH threshold for PA prepore-to-pore conversion.

A. Cytotoxicity assays of TEM8-expressing cells challenged with PA or PA-R344E (plus 1.9 nM FP59). The cells were treated with toxins in the absence or presence of 10 mM NH₄Cl for 3 h, then the toxins were removed and the cells were cultured in the presence of 10 mM NH₄Cl for 48 h and cell viability determined. NH₄Cl provided only slight protection. B. Analyses performed as in panel A showing that NH₄Cl can fully protect CMG2-expressing PR230 cells from PA plus FP59, but this protection is slightly less effective when the cells were treated with PA-R344E plus FP59, where the salt bridge between PA Arg-344 and CME2 Glu-122 is disrupted. C. Cytotoxicity assays done as above show that CMG2-E122K-expressing cells are more sensitive to PA-R344E than to native PA, due to restoration of the salt bridge, and this strengthened interaction restores high sensitivity to inhibition by NH₄Cl. D-F. Cytotoxicity assays of cells expressing CMG2 having the Tyr119 to His and His121 to Gln substitutions. The Tyr119 to His mutation makes cells largely insensitive to NH₄Cl protection when present alone (Panel D) or in combination with the His121 to Gln substitution (Panel F).