Cholesterol-binding cytolytic protein toxins

Abstract

Cholesterol-binding cytolysins (CBCs) are a large family of 50- to 60-kDa single-chain proteins produced by 23 taxonomically different species of Gram-positive bacteria from the genera Streptococcus, Bacillus, Clostridium, Listeria and Arcanobacterium. Apart pneumolysin, which is an intracytoplasmic toxin, all the other toxins are secreted in the extracellular medium. Among the species producing CBCs, only L. monocytogenes and L. ivanovii are intracellular pathogens which grow and release their toxins in the phagocytic cells of the host. CBCs are lethal to animals and highly lytic toward eukaryotic cells, including erythrocytes. Their lytic and lethal properties are suppressed by sulfhydryl-group-blocking agents and reversibly restored by thiols or other reducing agents. These properties are irreversibly abrogated by very low concentrations of cholesterol and other 3beta-hydroxysterols. Membrane cholesterol is thought to be the toxin-binding site at the surface of eukaryotic cells. Toxins molecules bind as monomers to the membrane surface with subsequent oligomerization into arc-and ring-shaped structures surrounding large pores generated by this process. Thirteen structural genes of the toxins (all chromosomal) have been cloned and sequenced to date. The deduced primary structure of the proteins shows obvious sequence homology particularly in the C-terminal part and a characteristic common consensus sequence containing a unique Cys residue (ECTGLAWEWWR) near the C-terminus of the molecules (except pyolysin and intermedilysin). However, another Cys residue outside this undecapeptide and closer to the C-terminus occurs in ivanolysin. Genetic replacement of the Cys residue in the consensus undecapeptide by certain amino acids demonstrated that this residue was not essential for toxin function. Other residues in the undecapeptide have been mutagenized, particularly the Trp residues. One of these Trp appeared critical for lytic activity. The recent elucidation of the 3-D structure of perfringolysin O provided interesting information on the structure-activity relationship. The molecule was divided into four domains. Three domains are arranged in a row, giving an elongated shape. Domain 3 is covalently connected to the N-terminal domain 1 and packed laterally against domain 2. Membrane interaction of the monomer appears to be mediated by domain 4, while, oligomerization involves several sites scattered throughout the sequence. The Trp-rich region around the conserved Cys residue within domain 4 is assumed to conformationally adapt to cholesterol, and domain 3 is envisaged to move across the "hinge" by which it is connected to domain 1.