title: The Cholesterol-dependent Cytolysin Membrane-binding Interface Discriminates Lipid Environments of Cholesterol to Support β-Barrel Pore Insertion.
abstract: The majority of cholesterol-dependent cytolysins (CDCs) utilize cholesterol as a membrane receptor, whereas a small number are restricted to the GPI-anchored protein CD59 for initial membrane recognition. Two cholesterol-binding CDCs, perfringolysin O (PFO) and streptolysin O (SLO), were found to exhibit strikingly different binding properties to cholesterol-rich natural and synthetic membranes. The structural basis for this difference was mapped to one of the loops (L3) in the membrane binding interface that help anchor the toxin monomers to the membrane after receptor (cholesterol) binding by the membrane insertion of its amino acid side chains. A single point mutation in this loop conferred the binding properties of SLO to PFO and vice versa. Our studies strongly suggest that changing the side chain structure of this loop alters its equilibrium between membrane-inserted and uninserted states, thereby affecting the overall binding affinity and total bound toxin. Previous studies have shown that the lipid environment of cholesterol has a dramatic effect on binding and activity. Combining this data with the results of our current studies on L3 suggests that the structure of this loop has evolved in the different CDCs to preferentially direct binding to cholesterol in different lipid environments. Finally, the efficiency of β-barrel pore formation was inversely correlated with the increased binding and affinity of the PFO L3 mutant, suggesting that selection of a compatible lipid environment impacts the efficiency of membrane insertion of the β-barrel pore.
authors.affiliation: Department of Microbiology and Immunology, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104.
authors.forename: Allison J
authors.initials: AJ
authors.lastname: Farrand
authors.affiliation: Department of Microbiology and Immunology, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104.
authors.forename: Eileen M
authors.initials: EM
authors.lastname: Hotze
authors.affiliation: Departments of Molecular and Cellular Medicine, Texas A&M University, College Station, Texas 77843.
authors.forename: Takehiro K
authors.initials: TK
authors.lastname: Sato
authors.affiliation: Department of Microbiology and Immunology, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104.
authors.forename: Kristin R
authors.initials: KR
authors.lastname: Wade
authors.affiliation: Department of Biochemistry and Molecular Biology, Tulane University School of Medicine, New Orleans, Louisiana 70112.
authors.forename: William C
authors.initials: WC
authors.lastname: Wimley
authors.affiliation: Departments of Molecular and Cellular Medicine, Texas A&M University, College Station, Texas 77843; Departments of Chemistry and Biochemistry and Biophysics, Texas A&M University, College Station, Texas 77843.
authors.forename: Arthur E
authors.initials: AE
authors.lastname: Johnson
authors.affiliation: Department of Microbiology and Immunology, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104. Electronic address: Rod-Tweten@ouhsc.edu.
authors.forename: Rodney K
authors.initials: RK
authors.lastname: Tweten
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D018407:Bacterial Physiological Phenomena; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D001427:Bacterial Toxins / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism / Q000382:microbiology*; D002784:Cholesterol / Q000378:metabolism*; D003603:Cytotoxins / Q000737:chemistry / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000737:chemistry / Q000378:metabolism*; D008081:Liposomes / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D011485:Protein Binding; D017433:Protein Structure, Secondary; D013301:Streptolysins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D003603:Cytotoxins; D006460:Hemolysin Proteins; D008081:Liposomes; D013301:Streptolysins; C025964:streptolysin O; C020452:Clostridium perfringens theta-toxin; D002784:Cholesterol
keywords: bacterial toxin; cholesterol; lipid structure; membrane lipid; membrane structure