LISTERIOLYSIN-O - CHOLESTEROL INHIBITS CYTOLYSIS BUT NOT BINDING TO CELLULAR MEMBRANES

Listeriolysin O (LLO) binds to cholesterol-containing membranes in whi ch it oligomerizes to form pores. Preincubation of the toxin with chol esterol is known to inhibit haemolysis, whereas the oxidized form of c holesterol has no inhibitory effect. Using immunoblot analyses and flo w cytometry we demonstrate that preincubation with cholesterol does no t influence binding of the listeriolysin-cholesterol complex to red bl ood cells, eukaryotic cells or artificial membranes. Lytic activity of membrane-bound LLO inactivated by cholesterol can be restored by enzy matic treatment with cholesterol oxidase. To determine the step at whi ch cholesterol inhibits lytic activity, we looked for pore formation u sing electron microscopy. Pores formed by purified listeriolysin could be directly visualized using erythrocyte ghosts. This property was lo st upon incubation of the toxin with cholesterol. Quantitative analysi s strongly suggest that inhibition of lysis by cholesterol is not due to decreased binding of listeriolysin to target membranes, but rather to an interference with a subsequent step leading to polymerization of the toxin.