Measuring kinetic drivers of pneumolysin pore structure

Journal article

Most membrane attack complex-perforin/cholesterol-dependent cytolysin (MACPF/CDC) proteins are thought to form pores in target membranes by assembling into pre-pore oligomers before undergoing a pre-pore to pore transition. Assembly during pore formation is into both full rings of subunits and incomplete rings (arcs). The balance between arcs and full rings is determined by a mechanism dependent on protein concentration in which arc pores arise due to kinetic trapping of the pre-pore forms by the depletion of free protein subunits during oligomerization. Here we describe the use of a kinetic assay to study pore formation in red blood cells by the MACPF/CDC pneumolysin from Streptococcus pneumoniae. We show that cell lysis displays two kinds of dependence on protein concentration. At lower concentrations, it is dependent on the pre-pore to pore transition of arc oligomers, which we show to be a cooperative process. At higher concentrations, it is dependent on the amount of pneumolysin bound to the membrane and reflects the affinity of the protein for its receptor, cholesterol. A lag occurs before cell lysis begins; this is dependent on oligomerization of pneumolysin. Kinetic dissection of cell lysis by pneumolysin demonstrates the capacity of MACPF/CDCs to generate pore-forming oligomeric structures of variable size with, most likely, different functional roles in biology.

Authors: Gilbert, RJC; Sonnen, AFP

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Springer
• Journal: European Biophysics Journal
• Volume: 45
• Issue: 4
• Pages: 365-376
• Publication date: 2016-02-23
• Acceptance date: 2015-12-07
• DOI: 10.1007/s00249-015-1106-x
• EISSN: 1432-1017
• ISSN: 0175-7571
• Language: English
• Keywords: pore formation; kinetics; MACPF/CDC; toroidal pore; oligomerization; membrane structure