INIBIOLP   05426
INSTITUTO DE INVESTIGACIONES BIOQUIMICAS DE LA PLATA "PROF. DR. RODOLFO R. BRENNER"
Unidad Ejecutora - UE
artículos
Título:
Relevance of fatty acid covalently bound to Escherichia coli alpha-Hemolysin and membrane microdomains in the oligomerization process
Autor/es:
VANESA HERLAX; SABINA MATÉ; OMAR RIMOLDI; LAURA BAKÁS
Revista:
JOURNAL OF BIOLOGICAL CHEMISTRY
Referencias:
Año: 2008 p. 1 - 1
ISSN:
0021-9258
Resumen:
a-hemolysin (HlyA) is an exotoxin secreted by some pathogenic strains of E.coli that causes lysis of several mammalian cells, including erythrocytes of different species. HlyA is synthesized as a protoxin: ProHlyA, which is activated by acylation at two internal lysines - K563 and K689. It has been proposed that pore formation is the mechanism of cytolytic activity for this toxin, as shown in experiments with whole cells, planar lipid membranes, and liposomes, but these experiments have yielded conflicting results about the structure of the pore. In this study, HlyA cystein-replacement mutant proteins of aminoacids have been labeled with Alexa-488 and Alexa-546. FRET measurements, employing labeled toxin bound to sheep ghost erythrocytes, have demonstrated that HlyA oligomerizes on erythrocytes membranes. As the cytotoxic activity is absolutely dependent on acylation, we have studied the role of acylation in the oligomerization, demonstrating that fatty acids are essential in this process.                                                                                                             On the other hand, FRET and the hemolytic activity decrease when the erythrocytes ghosts are cholesterol depleted, hence indicating the role of membrane microdomains in the clustering of HlyA. Simultaneously, HlyA was found in detergent- resistant membranes (DRMs). ProHlyA has also been found in DRMs, thus demonstrating that the importance of acyl chains in toxin oligomerization is the promotion of protein-protein interaction. These results change the concept of the main role assigned to acyl chain in the targeting of proteins to membrane microdomains.