title: Inactivation of the pore-forming toxin Sticholysin I by peroxynitrite: protection by cys groups incorporated in the toxin.
abstract: Sea anemones synthesize a variety of toxic peptides and proteins of biological interest. The Caribbean Sea anemone Stichodactyla helianthus, produces two pore-forming toxins, Sticholysin I (St I) and Stichloysin II (St II), with the ability to form oligomeric pores in cell and lipid bilayers characteristically lacking cysteine in their amino acid sequences. Recently, two mutants of a recombinant variant of Sticholysin I (rSt I) have been obtained with a Cys residue in functionally relevant regions for the pore-forming activity of the toxin: r St I F15C (in the amino terminal sequence) and r St I R52C (in the binding site). Aiming at characterizing the effects of oxidants in toxins devoid (r St I) or containing -SH moieties (r St I F15C and r St I R52C), we measured their hemolytic activity and pore forming capacity prior and after their incubation with peroxynitrite (ONOO(-)). At low ONOO(-)/Toxin ratios, nearly 0.8 Trp groups are modified by each added peroxynitrite molecule, and the toxin activity is reduced in ca. 20 %. On the other hand, in -SH bearing mutants only 0.5 Trp groups are modified by each peroxynitrite molecule and the toxin activity is only reduced in 10 %. The results indicated that Cys is the initial target of the oxidative damage and that Trp residues in Cys-containing toxins were less damaged than those in r St I. This relative protection of Trp groups correlates with a smaller loss of hemolytic activity and permeabilization ability in liposomes and emphasizes the relevance of Trp groups in the pore forming capacity of the toxins.
authors.affiliation: Department of Chemistry, Chemistry and Biology Faculty, Universidad de Santiago de Chile (USACH), Santiago, Chile, luis.leonm@usach.cl.
authors.forename: L
authors.initials: L
authors.lastname: León
authors.forename: E A
authors.initials: EA
authors.lastname: Lissi
authors.forename: G
authors.initials: G
authors.lastname: Celedón
authors.forename: G
authors.initials: G
authors.lastname: Gonzalez
authors.forename: F
authors.initials: F
authors.lastname: Pazos
authors.forename: C
authors.initials: C
authors.lastname: Alvarez
authors.forename: M E
authors.initials: ME
authors.lastname: Lanio
mesh_terms: D000328:Adult; D003545:Cysteine / Q000737:chemistry; D004912:Erythrocytes / Q000187:drug effects; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D008081:Liposomes; D009930:Organic Chemicals / Q000737:chemistry / Q000494:pharmacology; D010084:Oxidation-Reduction / Q000187:drug effects; D010539:Permeability; D030421:Peroxynitrous Acid / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D013438:Sulfhydryl Compounds; D055815:Young Adult
chemical_list: D008081:Liposomes; D009930:Organic Chemicals; D013438:Sulfhydryl Compounds; C121348:stycholysin I; D030421:Peroxynitrous Acid; D003545:Cysteine