title: Disentangling the roles of cholesterol and CD59 in intermedilysin pore formation.
abstract: The plasma membrane provides an essential barrier, shielding a cell from the pressures of its external environment. Pore-forming proteins, deployed by both hosts and pathogens alike, breach this barrier to lyse target cells. Intermedilysin is a cholesterol-dependent cytolysin that requires the human immune receptor CD59, in addition to cholesterol, to form giant β-barrel pores in host membranes. Here we integrate biochemical assays with electron microscopy and atomic force microscopy to distinguish the roles of these two receptors in mediating structural transitions of pore formation. CD59 is required for the specific coordination of intermedilysin (ILY) monomers and for triggering collapse of an oligomeric prepore. Movement of Domain 2 with respect to Domain 3 of ILY is essential for forming a late prepore intermediate that releases CD59, while the role of cholesterol may be limited to insertion of the transmembrane segments. Together these data define a structural timeline for ILY pore formation and suggest a mechanism that is relevant to understanding other pore-forming toxins that also require CD59.
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building, Imperial College London, London SW7 2AZ, UK.
authors.forename: Courtney M
authors.initials: CM
authors.lastname: Boyd
authors.affiliation: Department of Chemistry and Institute of Chemical Biology, Imperial College London, London SW7 2AZ, UK.|London Centre for Nanotechnology, University College London, London WC1H 0AH, UK.
authors.forename: Edward S
authors.initials: ES
authors.lastname: Parsons
authors.affiliation: MRC Centre for Transplantation, King's College London, 5th Floor Tower Wing, Guys' Hospital, London SE1 9RT, UK.
authors.forename: Richard A G
authors.initials: RA
authors.lastname: Smith
authors.affiliation: Department of Chemistry and Institute of Chemical Biology, Imperial College London, London SW7 2AZ, UK.
authors.forename: John M
authors.initials: JM
authors.lastname: Seddon
authors.affiliation: Department of Chemistry and Institute of Chemical Biology, Imperial College London, London SW7 2AZ, UK.
authors.forename: Oscar
authors.initials: O
authors.lastname: Ces
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building, Imperial College London, London SW7 2AZ, UK.
authors.forename: Doryen
authors.initials: D
authors.lastname: Bubeck
mesh_terms: D001430:Bacteriocins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D018957:CD59 Antigens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D002784:Cholesterol / Q000737:chemistry / Q000378:metabolism*; D054884:Host-Pathogen Interactions; D006801:Humans; D018625:Microscopy, Atomic Force; D018272:Porins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000072417:Protein Domains / Q000235:genetics
chemical_list: D001430:Bacteriocins; D018957:CD59 Antigens; D018272:Porins; C403910:intermedilysin protein, Streptococcus intermedius; C051590:CD59 protein, human; D002784:Cholesterol