E. coli &lt;FONT FACE=Symbol&gt;a&lt;/FONT&gt;-hemolysin: a membrane-active protein toxin

Goñi, Félix M.; Ostolaza, Helena

doi:10.1590/s0100-879x1998000800002

Cited by 29 publications

(15 citation statements)

References 101 publications

(61 reference statements)

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“…It is a member of the so-called "RTX family," a group of proteins characterized by the presence of a Gly-and Asp-rich nonapeptide sequence repeated in tandem near the protein C terminus (for reviews, see Refs. [1][2][3][4][5]. These repeats constitute a Ca 2ϩ -binding domain whose structure has been solved at high resolution for a non-toxin member of the RTX family, the alkaline protease from Pseudomonas aeruginosa (6).…”

Section: ␣-Hemolysinmentioning

confidence: 99%

A Receptor-binding Region in Escherichia coli α-Haemolysin

Cortajarena¹,

Goñi

Ostolaza

2003

Journal of Biological Chemistry

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Escherichia coli ␣-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., Goñ i, 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⌬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. 1 is a 107-kDa protein toxin secreted by pathogenic strains of Escherichia coli. It is a member of the so-called "RTX family," a group of proteins characterized by the presence of a Gly-and Asp-rich nonapeptide sequence repeated in tandem near the protein C terminus (for reviews, see Refs. 1-5). These repeats constitute a Ca 2ϩ -binding domain whose structure has been solved at high resolution for a non-toxin member of the RTX family, the alkaline protease from Pseudomonas aeruginosa (6). HlyA first binds a receptor on the cell surface, a ␤ 2 -integrin in leukocytes (7) or glycophorin in red blood cells (8), and then becomes inserted in the cell membrane. Recent data indicate that insertion may take place in the absence of Ca 2ϩ (9, 10), but Ca 2ϩ binding to the nonapeptide repeat domain is essential for membrane lysis (9,11,12). Note that the Ca 2ϩ -binding domain is located near the protein C terminus, whereas the membrane insertion domain is located near the N terminus (9, 10, 13). The present study is devoted to exploring the early stages of HlyA interaction with the target cell, namely its binding to the surface receptor. In particular, our investigation is aimed at the region(s) of the protein that bind(s) the receptor glycophorin on mammalian erythrocytes (8). A number of previous studies suggest that a region between the repeat domain and the C terminus may be involved in binding this specific receptor. For example, Bejerano et al. (14) have described two amino acid "blocks," located after the nonapeptide repeats, in the C terminus of the adenylate cyclase toxin (another member of the RTX family). Block A (15 amino acids) is essential for the toxic activity since it is required for the toxin binding and insertion into the membrane. Deletion of block B, however, does not affect the toxin activity. HlyA possesses homologous A and B blocks (Fig. 1), and it has been shown (14) that deletion of a region that includes the last two residues of block A, the connection between the blocks, and the first nine amino acids of block B...

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Section: ␣-Hemolysinmentioning

confidence: 99%

A Receptor-binding Region in Escherichia coli α-Haemolysin

Cortajarena¹,

Goñi

Ostolaza

2003

Journal of Biological Chemistry

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“…RTX toxins are proteins secreted by type I export systems that destroy target cells by disrupting cell membranes [19]. Their protein structure contains a domain of glycine-rich nonapeptide calcium binding repeats, which are required for the toxin’s activity [20, 21]. The toxins are activated by covalent fatty acid acylation of specific lysines, which represent a unique characteristic of this group of toxins [22–24].…”

Section: Introductionmentioning

confidence: 99%

Pore forming activity of the potent RTX-toxin produced by pediatric pathogen Kingella kingae: Characterization and comparison to other RTX-family members

Bárcena-Uribarri

Benz

Winterhalter

et al. 2015

Biochimica et Biophysica Acta (BBA) - Biomembranes

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Pediatric septic arthritis in patients under age of four is frequently caused by the oral Gram-negative bacterium Kingella kingae. This organism may be responsible for a severe form of infective endocarditis in otherwise healthy children and adults. A major virulence factor of K. kingae is RtxA, a toxin that belongs to the RTX (Repeats-in-ToXin) group of secreted pore forming toxins. To understand the RtxA effects on host cell membranes, the toxin activity was studied using planar lipid bilayers. K. kingae strain PYKK081 and its isogenic RtxA-deficient strain, KKNB100, were tested for their ability to form pores in artificial membranes of asolectin/n-decane. RtxA, purified from PYKK081, was able to rapidly form pores with an apparent diameter of 1.9 nm as measured by the partition of nonelectrolytes in the pores. The RtxA channels are cation-selective and showed strong voltage-dependent gating. In contrast to supernatants of PYKK081, those of KKNB100 did not show any pore forming activity. We concluded that RtxA toxin is the only secreted protein from K. kingae forming large channels in host cell membranes where it induces cation flux leading to programmed cell death. Furthermore, our findings suggested that the planar lipid bilayer technique can effectively be used to test possible inhibitors of RTX toxin activity and to investigate the mechanism of the toxin binding to the membrane.

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“…Their members must undergo a transformation from a water-soluble to an insoluble, membrane-bound form to exert their effect on the target cell (for reviews, see Refs. [2][3][4][5].…”

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confidence: 99%