title: Asp-863 is a key residue for calcium-dependent activity of Escherichia coli RTX toxin alpha-haemolysin.
abstract: alpha-Haemolysin is a protein toxin secreted by pathogenic strains of Escherichia coli and requires sub-millimolar Ca(2+) for optimum lytic activity. As a member of the so-called RTX toxin family it contains a Gly-rich, Asp-rich Ca(2+)-binding domain, consisting of a series of nonapeptides repeated in tandem. Asp-863 is located immediately after the last-but-one nonapeptide. A mutant in which Asp-863 has been substituted by Gly displays a requirement for Ca(2+) that is 100-fold higher than the wild-type. Membrane lytic activity, as well as a conformational change revealed through an increase in intrinsic fluorescence, and the appearance of Ca(2+)-bound protein monomers resolvable by fast protein liquid chromatography, are all three dependent on Ca(2+) concentrations in the 2-20 mM range. Most RTX toxins have an Asp or Glu residue located at a position homologous to Asp-863, thus the key role of this residue for Ca(2+) requirements of alpha-haemolysin may be a general feature of this family of toxins.
authors.affiliation: Unidad de Biofísica (Centro Mixto CSIC-UPV/EHU), and Departamento de Bioquímica, Universidad del País Vasco, Aptdo 644, 48080 Bilbao, Spain.
authors.forename: Aitziber L
authors.initials: AL
authors.lastname: Cortajarena
authors.forename: Félix M
authors.initials: FM
authors.lastname: Goñi
authors.forename: Helena
authors.initials: H
authors.lastname: Ostolaza
mesh_terms: D000595:Amino Acid Sequence; D001224:Aspartic Acid / Q000378:metabolism*; D001427:Bacterial Toxins / Q000737:chemistry / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002850:Chromatography, Gel; D004926:Escherichia coli / Q000378:metabolism*; D005453:Fluorescence; D006460:Hemolysin Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid
chemical_list: D001427:Bacterial Toxins; D006460:Hemolysin Proteins; D001224:Aspartic Acid; D002118:Calcium