Perfringolysin O Structure and Mechanism of Pore Formation as a Paradigm for Cholesterol-Dependent Cytolysins

Abstract

Cholesterol-dependent cytolysins (CDCs) constitute a family of pore forming toxins secreted by Gram positive bacteria. These toxins form transmembrane pores by inserting a large β-barrel into cholesterol-containing membrane bilayers. Binding of water-soluble CDCs to the membrane triggers the formation of oligomers containing 35-50 monomers. The coordinated insertion of more than seventy β-hairpins into the membrane requires multiple structural conformational changes. Perfringolysin O (PFO), secreted by Clostridium perfringens, has become the prototype for the CDCs. In this chapter, we will describe current knowledge on the mechanism of PFO cytolysis, with special focus on cholesterol recognition, oligomerization, and the conformational changes involved in pore formation.

Perfringolysin O Theta Toxin as a Tool to Monitor the Distribution and Inhomogeneity of Cholesterol in Cellular Membranes

“…Intermedilysin is an exception to the cholesterol requirement as intermedilysin makes use of CD59 for binding to the plasma membrane [ 4 ]. CDCs are secreted by a variety of Gram-positive bacteria including but not limited to Bacillus , Listeria , Streptococcus , Clostridium , and Lactobacillus [ 5 ]. Binding of these water-soluble CDCs to cholesterol in the exofacial leaflets of the plasma membrane induces the oligomerization of the toxins (30–50 monomers) [ 1 , 6 , 7 , 8 ].…”

Atomic force microscopy of membrane pore formation by cholesterol dependent cytolysins

“…The structure of the water-soluble monomeric form has been solved for several CDCs by X-ray crystallography ( Figure 1A, [12][13][14][15][16][17][18][19]). It appears to be well conserved through the CDC family, consists of at least 30 protein members [20], and shares the pore-forming topology with the large membrane attack complex/perforin superfamily [6]. CDC monomers comprise four domains, which are responsible for membrane binding, oligomerization, and membrane insertion.…”

Transverse distribution of plasma membrane bilayer cholesterol: Picking sides

“…An early study by Bittman et al used the kinetics of filipin association to infer that cholesterol was in excess in the outer leaflet of the plasma membrane of Mycoplasma gallisepticum . (This technique is described in a later section.…”

“…PFO and its derivatives do not bind uniformly to the cholesterol present in membranes and therefore do not report its quantity directly. Rather, these toxins only associate with the available (accessible or chemically active) fraction of the cholesterol, because the bulk of the sterol is complexed stoichiometrically with the bilayer phospholipids and is thus competitively inhibited from binding the probe . The membrane cholesterol concentration dependence of probe binding therefore has a threshold that reflects the relative affinities of the phospholipids and the probes for the sterol .…”

“…Rather, these toxins only associate with the available (accessible or chemically active) fraction of the cholesterol, because the bulk of the sterol is complexed stoichiometrically with the bilayer phospholipids and is thus competitively inhibited from binding the probe. 22,[57][58][59] The membrane cholesterol concentration dependence of probe binding therefore has a threshold that reflects the relative affinities of the phospholipids and the probes for the sterol. [60][61][62] Calibration with synthetic vesicles of the appropriate phospholipid composition is therefore critical.…”