Search results for "Hemolysi"
showing 10 items of 241 documents
Staphylococcal alpha-toxin: repair of a calcium-impermeable pore in the target cell membrane
2000
Staphylococcal alpha-toxin forms heptameric pores that render membranes permeable for monovalent cations. The pore is formed by an amphipathic beta-barrel encompassing amino acid residues 118-140 of each subunit of the oligomer. Human fibroblasts are susceptible to alpha-toxin but are able to repair the membrane lesions. Thereby, toxin oligomers remain embedded in the plasma membrane and exposed to the extracellular medium. In this study, we sought to detect structural changes occurring in the pore-forming sequence during lesion repair. Single cysteine substitution mutants were labelled with the environmentally sensitive fluorochrome acrylodan and, after mixing with wild-type toxin, incorpo…
Purification and characterization of a pore-forming protein from the marine sponge Tethya lyncurium
1992
A pore-forming protein was detected and purified for the first time from a marine sponge (Tethya lyncurium). The purified protein has a polypeptide molecular mass of 21 kDa and a pI of 6.4. Tethya pore-forming protein (also called Tethya hemolysin) rapidly lysed erythrocytes from a variety of organisms. After binding to target membranes, the hemolysin resisted elution with EDTA, salt or solutions of low ionic strength and hence resembled an integral membrane protein. Erythrocytes could be protected from hemolysis induced by Tethya hemolysin by addition of 30 mM dextran 4 (4-6 kDa; equivalent hydrodynamic diffusion radius, 1.75-2.3 nm) to the extracellular medium, but not by addition of unch…
Toxicity and mode of action of Bacillus thuringiensis Cry proteins in the Mediterranean corn borer, Sesamia nonagrioides (Lefebvre)
2006
ABSTRACT Sesamia nonagrioides is one of the most damaging pests of corn in Spain and other Mediterranean countries. Bt corn expressing the Bacillus thuringiensis Cry1Ab toxin is being grown on about 58,000 ha in Spain. Here we studied the mode of action of this Cry protein on S. nonagrioides (binding to specific receptors, stability of binding, and pore formation) and the modes of action of other Cry proteins that were found to be active in this work (Cry1Ac, Cry1Ca, and Cry1Fa). Binding assays were performed with 125 I- or biotin-labeled toxins and larval brush border membrane vesicles (BBMV). Competition experiments indicated that these toxins bind specifically and that Cry1Aa, Cry1Ab, an…
Superoxide generation by human neutrophils induced by low doses of Escherichia coli hemolysin.
1991
Escherichia coli hemolysin (Hly) was isolated from bacterial culture supernatants by polyethylene glycol precipitation and centrifugation in glycerol density gradients. The toxin preparations contained less than 1 mol of lipopolysaccharide per 10 mol of protein, and they had no fatty acids. The capacity of purified hemolysin to stimulate superoxide anion production in polymorphonuclear leukocytes was monitored kinetically in a lumimeter by using the lucigenin assay and was correlated with the kinetics of transmembrane pore formation. When applied to leukocytes suspended in protein-free buffer, very low concentrations (0.02 to 0.1 HU/ml) of the toxin strongly stimulated the production of sup…
Hypersusceptibility of neutrophil granulocytes towards lethal action of free fatty acids contained in enzyme-modified atherogenic low density lipopro…
2008
Abstract Objective The bulk of LDL entrapped in the arterial intima is modified by hydrolytic enzymes, leading to extensive cleavage of cholesterylesters and liberation of fatty acids. The latter induce apoptosis in endothelial cells but are far less cytotoxic towards macrophages. We have compared the cytotoxic effects of enzymatically modified LDL (E-LDL) on macrophages and polymorphonuclear granulocytes (PMN). Methods and results E-LDL displayed toxicity towards PMN at far lower concentrations than towards monocyte-derived macrophages. Native or oxidized LDL had no effect. Free fatty acids contained in E-LDL were the cause of the observed toxicity, which could be mimicked by linoleic acid…
Effect of EDTA and citrate on the functional activity of the first component of complement, C1, and the C1q subcomponent.
1985
The first component of complement, C1, is a calcium-dependent complex of the three distinct subcomponents, C1q, C1r, and C1s. Earlier observations revealed that treatment of C1 with EDTA led to a loss of hemolytic C1 activity even after recalcification. Therefore, it was of interest to study whether EDTA has an additional effect on C1 and its subcomponents, beside its chelating capacity. The chelating effect of EDTA was compared to that of citrate. It was found that treatment of C1 or C1 with EDTA followed by addition of Ca++ led to a loss of hemolytic activity up to 90%, depending on EDTA concentration. Even pretreatment of EDTA with varying amounts of Ca++ did not prevent the inactivation…
The cytotoxin-hemolysin genes of human and eel pathogenic Vibrio vulnificus strains: comparison of nucleotide sequences and application to the geneti…
2005
Vibrio vulnificus can be divided into two groups on the basis of pathogenesis. Group 1 is pathogenic only to humans, whereas group 2 is pathogenic to eels and occasionally to humans. Although both groups produce a 50-kDa cytotoxin-hemolysin (V. vulnificus hemolysin; VVH), the toxins are different. In the present study, the nucleotide sequence of the toxin gene (vvhA ) of strain CDC B3547 (a group 2 strain) was determined, and the deduced amino acid sequence was compared to that of strain L-180 (a group 1 strain). The nucleotide sequence of vvhA of strain CDC B3547 was about 96% identical with that of strain L-180, which results in a difference of 3 amino acid residues in the C-terminal lect…
Cholesterol reporter molecules.
2007
Cholesterol is a major constituent of the membranes in most eukaryotic cells where it fulfills multiple functions. Cholesterol regulates the physical state of the phospholipid bilayer, affects the activity of several membrane proteins, and is the precursor for steroid hormones and bile acids. Cholesterol plays a crucial role in the formation of membrane microdomains such as “lipid rafts” and caveolae. However, our current understanding on the membrane organization, intracellular distribution and trafficking of cholesterol is rather poor. This is mainly due to inherent difficulties to label and track this small lipid. In this review, we describe different approaches to detect cholesterol in …
Structural characterisation of the natural membrane-bound state of melittin: a fluorescence study of a dansylated analogue
1997
Abstract The binding of a dansylated analogue of melittin (DNC–melittin) to natural membranes is described. The cytolytic peptide from honey bee venom melittin was enzymatically labelled in its glutamine-25 with the fluorescent probe monodansylcadaverine using guinea pig liver transglutaminase. The labelled peptide was characterised functionally in cytolytic assays, and spectroscopically by circular dichroism and fluorescence. The behaviour of DNC–melittin was, in all respects, indistinguishable from that of the naturally occurring peptide. We used resonance energy transfer to measure the state of aggregation of melittin on the membrane plane in synthetic and natural lipid bilayers. When bo…
Interaction ofEscherichia colihemolysin with biological membranes
2001
Escherichia coli hemolysin (HlyA) is a membrane-permeabilizing protein belonging to the family of RTX-toxins. Lytic activity depends on binding of Ca2(+) to the C-terminus of the molecule. The N-terminus of HlyA harbors hydrophobic sequences that are believed to constitute the membrane-inserting domain. In this study, 13 HlyA cysteine-replacement mutants were constructed and labeled with the polarity-sensitive fluorescent probe 6-bromoacetyl-2-dimethylaminonaphthalene (badan). The fluorescence emission of the label was examined in soluble and membrane-bound toxin. Binding effected a major blue shift in the emission of six residues within the N-terminal hydrophobic domain, indicating inserti…