| The Cholesterol-Dependent Cytolysins (CDCs) are a large family of pore-forming toxins, which share 40-70% primary structure homology and exhibit a four domain (D1-D4) tertiary structure. Two classes of CDCs currently exist, intermedilysin (ILY) and perfringolysin O (PFO)-like CDCs. The pore-forming mechanism of both classes of CDCs is absolutely dependent on membrane cholesterol. However, each class differs in their initial contact with the cell surface. PFO and most other CDCs can bind directly to cholesterol-rich membranes, and it is thought that cholesterol is the cell surface receptor. For ILY, it initially interacts with the membrane via a different receptor, human CD59 (huCD59). These observations presented an enigma; how did cholesterol contribute to the mechanism of PFO and ILY if cholesterol was the receptor for one and not the other? To resolve this problem a detailed investigation was performed on the interaction of both toxins with the membrane surface. In these studies we showed that the ILY D4 loops, which consist of the three short, hydrophobic loops (L1-L3) and the conserved undecapeptide, insert into the membrane, similar to PFO, but only after ILY has bound to its receptor. Furthermore, the insertion of these loops is necessary for the subsequent formation of the ILY oligomeric prepore and pore structures. These results led to the structural basis for the cholesterol dependence of the CDCs, a problem that has been unresolved for over 50 years. We have shown the membrane insertion of the L1-L3 loops is the basis for the cholesterol-dependence of both classes of CDCs. They do not insert into membranes that lack or are substantially depleted of cholesterol. It is the L1-L3 loops that mediate the interaction between the CDCs and cholesterol-rich membranes. Therefore, these studies have resolved the basis for the cholesterol dependence of both ILY and PFO-like CDCs. |
