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Lysis
About: Lysis is a research topic. Over the lifetime, 6072 publications have been published within this topic receiving 216978 citations.
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TL;DR: An initial proteomic map of membrane and cell wall associated proteins for S. aureus is presented and it is demonstrated that cell lysis with lysostaphin, followed by solubilization with urea, thiourea, amidosulfobetaine 14 and dithiothreitol is an effective method, yielding a sample comprising proteins of wide molecular ranges and isoelectric points with minimum contamination from cytosolic proteins.
Abstract: Pathogenesis of Staphylococcus aureus, an opportunistic human pathogen, is complex and involves many virulence factors including an array of surface proteins (adhesins) that promote bacterial interactions with extracellular matrix components A better understanding of these interactions can be achieved by studying the expression of membrane and cell wall associated proteins using a proteome analysis approach To accomplish this, our goal here was to construct a reference map of membrane and cell wall associated proteins for S aureus Various lytic and solubilization methods have been tested to identify a suitable methodology for detection of these proteins in two-dimensional electrophoresis (2DE) Results demonstrate that cell lysis with lysostaphin, which lyses staphylococcal peptidoglycan, followed by solubilization with urea, thiourea, amidosulfobetaine 14 (ASB 14) and dithiothreitol (DTT) is an effective method, yielding a sample comprising proteins of wide molecular ranges and isoelectric points with minimum contamination from cytosolic proteins Mass spectrometric analysis was employed to identify the membrane and cell surface proteins present in the sample and consequently an initial proteomic map of membrane and cell wall associated proteins for S aureus is presented
93 citations
TL;DR: It was demonstrated that during reconstitution the symmetric vesicles were formed at 19 mM free equilibrium octyl glucoside by the association of the 6-S protein complexes with the phospholipids, and the asymmetric vESicles were form at 10--16 mM octyl glucose when the 19-S complexes associated with the lipids.
Abstract: The dissociation and reconstitution of the Semliki Forest virus membrane using the nonionic detergent octyl beta-D-glucoside was studied by sucrose density gradient centrifugation. The dissociation occurred in three stages: lysis at a free equilibrium octyl glucoside concentration of 14--18 mM, solubilization at 18--20 mM, and delipidation of the spike glycoproteins at the critical micellar concentration (22 mM) or higher. After solubilization the spike glycoproteins were present as soluble complexes with sedimentation coefficients of 19 S and 6 S. The 6-S form probably corresponded to a glycoprotein monomer complexed to detergent and the 19-S form consisted of oligomeric detergent-protein complexes. The two forms were in slow equilibrium with each other. When the soluble spike protein complexes and egg lecithin solubilized with octyl glucoside were mixed and the octyl glucoside concentration lowered either by dialysis or by dilution, reconstitution occurred. Three types of products were obtained: vesicles with 30% of the spike protein facing inwards and 70% facing outwards, vesicles with virtually all (95%) of the spike proteins pointing outwards, and small protein-rich soluble aggregates [Helenius et al. (1977) J. Cell Biol. 75, 866]. It was demonstrated that during reconstitution the symmetric vesicles were formed at 19 mM free equilibrium octyl glucoside by the association of the 6-S protein complexes with the phospholipids, and the asymmetric vesicles were formed at 10--16 mM octyl glucoside when the 19-S complexes associated with the lipids. Asymmetric membrane vesicles were also obtained when membrane penicillinase from Bacillus licheniformis was reconstituted with egg lecithin using octyl glucoside. It could be shown that the penicillinase was oligomeric at the octyl glycoside concentration where the reconstitution occurred. The results demonstrate that different mechanisms of reconstitution give rise to the symmetric and the asymmetric vesicles. The critical factor in determining the mechanism is the state of aggregation of the proteins at the octyl glucoside concentration where membranes begin to form from the solubilized lipids.
93 citations
TL;DR: This system realizes timely cell disruption immediately after the PHB content in the cell reaches a maximum level and spontaneously induced self-disruption was spontaneously induced, releasing intracellularly accumulated PHB into the culture broth.
Abstract: In order to establish a novel recovery system for polyhydroxyalkanoates, a self-disruptive strain of Bacillus megaterium that responds to substrate exhaustion was constructed. A gene cassette carrying the lysis system of Bacillus amyloliquefaciens phage – holin and endolysin – was inserted into the Escherichia coli-Bacillus subtilis shuttle vector pX under the control of a xylose-inducible expression system, xylR-xylA
′. In this system, the expression of a target gene is induced by xylose but inhibited by glucose, which acts as an anti-inducer. B. megaterium was transformed with pX conveying the phage lysis system, which was integrated into the amyE locus of chromosomal DNA of B. megaterium by homologous recombination. The lysis system caused self-disruption of the transformant cells effectively even when expression of the lysis genes was induced during stationary phase. For the production of polyhydroxybutyrate (PHB), the transformant was grown in a medium containing glucose as a substrate in the presence of xylose. When the glucose concentration approached zero, self-disruption was spontaneously induced, releasing intracellularly accumulated PHB into the culture broth. This system realizes timely cell disruption immediately after the PHB content in the cell reaches a maximum level.
93 citations
TL;DR: A mutant of phage T4 has been found which permits lysis of infected bacteria in the absence of lysozyme, and the new mutation may affect a phage-directed synthesis of bacterial cell wall components, hence the name spackle.
92 citations
TL;DR: It is shown that, after interaction of APOL1 with lipid bilayers at acidic pH, subsequent pH neutralization triggers the opening of pH-gated channels that selectively conduct cations across the bilayer, which is consistent with a model of lysis that involves endocytic recycling of APol1 and the formation of cation-selective channels, at neutral pH, in the parasite plasma membrane.
Abstract: Apolipoprotein L-1 (APOL1), the trypanolytic factor of human serum, can lyse several African trypanosome species including Trypanosoma brucei brucei, but not the human-infective pathogens T. brucei rhodesiense and T. brucei gambiense, which are resistant to lysis by human serum. Lysis follows the uptake of APOL1 into acidic endosomes and is apparently caused by colloid-osmotic swelling due to an increased ion permeability of the plasma membrane. Here we demonstrate that nanogram quantities of full-length recombinant APOL1 induce ideally cation-selective macroscopic conductances in planar lipid bilayers. The conductances were highly sensitive to pH: their induction required acidic pH (pH 5.3), but their magnitude could be increased 3,000-fold upon alkalinization of the milieu (pKa = 7.1). We show that this phenomenon can be attributed to the association of APOL1 with the bilayer at acidic pH, followed by the opening of APOL1-induced cation-selective channels upon pH neutralization. Furthermore, the conductance increase at neutral pH (but not membrane association at acidic pH) was prevented by the interaction of APOL1 with the serum resistance-associated protein, which is produced by T. brucei rhodesiense and prevents trypanosome lysis by APOL1. These data are consistent with a model of lysis that involves endocytic recycling of APOL1 and the formation of cation-selective channels, at neutral pH, in the parasite plasma membrane.
92 citations