This model would require large conformational changes in domain 2 along with the breaking of many hydrogen bonds as the Greek-key motif peels away from the center of the domain. Petosa et al. proposed a model of how this occurs. Insertion of the PA Greek key motifs into the membrane occurs when the heptamer is acidified. On artificial bilayers, this occurs when the pH is dropped from 7.4 to 6.5, suggesting that the trigger for insertion involves a titration of histidines. This indeed fits the sequence of PA since domain 2 contains a number of histidines (shown as asterisks in figure 9a). Three histidine residues are found in the disordered loop, one of which lies with a Greek-key histidine within a cluster of polar amino acids. This cluster (including the two histidines, three arginines and one glutamate) is embedded at the top of the Greek-key motif, so it is easy to see that the protonation of these histidines would disrupt the cluster. Furthermore, another histidine is located at the base of the Greek-key motif along with a number of hydrophobic residues (on the green segment in figures 7 and 9a). At pH 7.4 this segment is ordered, but when the crystals are grown at pH 6.0, it becomes disordered. This order to disorder transition is the initial step of PA membrane insertion.
