Abstract
Patch clamping was used to investigate, for the first time, the electrophysiological properties of pores formed by a cholesterol-dependent cytolysin in the membrane of nucleated cells rather than in artificial membranes as previously. Pneumolysin pores in inside-out patches of CHO cell membranes had a broad range of conductance classified into small (<200 pS), medium (>200 pS and <1 nS) and large (>1 nS). A lytic-deficient mutant of pneumolysin (W433F) induced a similar spectrum of channels but there were more small channels than with wild-type (WT) toxin and the number of events was decreased. The WT toxin also induced channels when given to the inside surface of the plasma membrane.
Highlights
The pneumococcal toxin pneumolysin belongs to the family of cholesterol dependent cytolysins (CDCs)
We first tested the ability of pneumolysin to form pores when applied to the outside surface of Chinese Hamster Ovary (CHO)
Because of the complex behaviour and multiple conductances of pneumolysin-induced channels, it was not possible to obtain a quantitative measure of the open-time distribution of the various conductance states
Summary
The pneumococcal toxin pneumolysin belongs to the family of cholesterol dependent cytolysins (CDCs) These toxins bind to cholesterol-containing membrane, oligomerise and generate large pores that disrupt the membrane integrity, leading to cell death [1]. The patch-clamp technique was used in inside out and outside out configurations to study, for the first time, the pores of one of the CDCs (pneumolysin) across the membrane of a nucleated cell. In these experiments, we studied the lytic-deficient pneumolysin mutant, W433F, to investigate if lytic deficiency correlated with absence of electrophysiological properties. Trp433 is in a highly conserved region in all the CDCs is involved in the binding of the toxin monomers to the cell membrane [4]
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have