A receptor-binding region in Escherichia coli alpha-haemolysin.
Escherichia coli alpha-hemolysin (HlyA) is a 107-kDa protein toxin with a wide range of mammalian target cells. Previous work has shown that glycophorin is a specific receptor for HlyA in red blood cells (Cortajarena, A. L., Goñi, F. M., and Ostolaza, H. (2001) J. Biol. Chem. 276, 12513-12519). The present study was aimed at identifying the glycophorin-binding region in the toxin. Data in the literature pointed to a short amino acid sequence near the C terminus as a putative receptor-binding domain. Previous sequence analyses of several homologous toxins that belong, like HlyA, to the so-called RTX toxin family revealed a conserved region that corresponded to residues 914-936 of HlyA. We therefore prepared a deletion mutant lacking these residues (HlyA Delta 914-936) and found that its hemolytic activity was decreased by 10,000-fold with respect to the wild type. This deletion mutant was virtually unable to bind human and horse red blood cells or to bind pure glycophorin in an affinity column. The peptide Trp914-Arg936 had no lytic activity of its own, but it could bind glycophorin reconstituted in lipid vesicles. Moreover, the peptide Trp914-Arg936 protected red blood cells from hemolysis induced by wild type HlyA. It was concluded that amino acid residues 914-936 constitute a major receptor-binding region in alpha-hemolysin.
Cortajarena
Aitziber L
AL
Unidad de Biofísica, Consejo Superior de Investigaciones Científicas-Euskal Herriko Unibertsitatea and Departamento de Bioquímica, Universidad del País Vasco, Aptdo. 644, 48080 Bilbao, Spain.
Goni
Félix M
FM
Ostolaza
Helena
H
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D017124:Conserved Sequence; D004912:Erythrocytes / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017353:Gene Deletion; D006021:Glycophorins / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006461:Hemolysis; D006736:Horses / Q000097:blood; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010446:Peptide Fragments / Q000737:chemistry; D012091:Repetitive Sequences, Nucleic Acid; D016415:Sequence Alignment; D013329:Structure-Activity Relationship
D029968:Escherichia coli Proteins; D006021:Glycophorins; D006460:Hemolysin Proteins; C036507:Hlya protein, E coli; D010446:Peptide Fragments