Permeabilization Membranes

Adtveoitages. The antibody technology is remarkably convenient. It requires only that a significant fraction of FLPEP be receptor bound ( 10%). The assay can be set up in a few minutes and is applicable to membranes, permeabilized cells, cells, or any other preparation in which the receptor concentration is in the range of 1 niV or greater. We use this assay routinely to analyze ligand dissociation kinetics. 

Some of the compounds have been evaluated for their ability to induce mitochondrial membrane permeabilization, which is believed to play a fundamental role in the regulation of programmed cell apoptosis. The effects observed are not in line with a conventional model of action related to the accumulation of the lipophilic compounds in the cell, and further studies are necessary.281... 

Ruden S, Hilpert K, Berditsch M, Wadhwani P, Ulrich AS (2009) Synergistic interaction between silver nanoparticles and membrane-permeabilizing antimicrobial peptides. Antimicrob Agents Chemother 53 3538-3540... 

Knorr, D., Heinz, V., Angersbach, A., and Lee, D.-U. 2002. Membrane permeabilization and inactivation mechanisms of biological systems by emerging technologies. In Engineering and Food for the 21st Century (J. Welti-Chanes, G.V. Barbosa-Canovas, and J.M. Aguilera, eds), pp. 699-713. CRC Press, Boca Raton, FL. 

Although there is evidence to support all of the above models, it has been suggested that membrane permeabilization by host defense peptides likely results from elements of all the proposed models. ... 

Defensins Mammals, birds, Invertebrates, plants, fungi Membrane permeabilization, macromolecular synthesis inhibition Cell proliferation/differentiation chemotaxis induction of gene expression adaptive immune polarization in vivo protection cytokine/chemokine induction CCR6, TLRs -1, -2 and -4... 

Alcohols and acetone are the simplest kinds of membrane permeabilization agents. They act by dissolving membrane lipids, thus rendering the membrane permeable to antibodies (5). Because of their coagulant effects on proteins, these solvents can be used as a one-step fixative and permeant (18). 

B17. Bussing, A., Vervecken, W, Wagner, M., Wagner, B., Pfuller, U., and Schietzel, M., Expression of mitochondrial Apo2.7 molecules and caspase-3 activation in human lymphocytes treated with the ribosome-inhibiting mistletoe lectins and the cell membrane permeabilizing viscotoxins. Cytometry 37, 133-139 (1999). 

Cesari et al. (2007) demonstrated on bovine and human sperm in vitro that potato aspartic proteases 1 and 3 are effective spermicidal compounds. Spermicidal activity is due to membrane permeabilization and is not related to the proteolytic activity. Cesari et al. (2007) suggest vaginal contraception as a possible applieation. 

Busse H-J, Wostmann C, Bakker EP The bactericidal action of streptomycin Membrane permeabilization caused by the insertion of mistranslated proteins into the cytoplasmic membrane of Escherichia coli and subsequent caging of the antibiotic inside the cells due to degradation of these proteins. J Gen Microbiol 1992 138 551. [PMID 1375623]... 

Armstrong JS. Mitochondrial Membrane Permeabilization the sine qua non for cell death. Bio Essays 2006 28 252-260. 

The limitations of the application of conventional detergents mentioned above can be circumvented by replacing this approach with cell membrane permeabilization by microwave heating. Improved detection of intracellular antigens can be obtained with microwave heating used in combination with flow cytometry. This approach yields histogram patterns that show clear discrimination between intact cells and cell debris (Fig. 9.5). 

Membrane permeabilization activity of peptides is currently measured by the use of artificial membrane bilayers, such as liposomes or erythrocytes. The hposome leakage assay can be performed by using spectrofluorimetry with a concentration-dependent quenching of a dye (calcein, carboxyfluorescein) encapsulated in liposomes. Disruption of hposomes in the presence of peptide-inducing leakage will lead to an increase in the fluorescence intensity of the liposome solution. Erythrocyte lysis assay is based on the absorption of hemoglobin, which can be measured once released into the extracellular medium upon erythrocyte lysis in the presence of peptide. 

Membrane destabilization can be simultaneously monitored by using propidium iodide. This non fluorescent and poorly permeant small molecule enters into the cells only upon membrane permeabilization/ destabilization and becomes fluorescent upon binding to DNA in the nucleus. In the presence of melittin, cells are also rapidly (200 s) red-labeled upon peptide induced membrane permeabilization (Figure 16.4D). It can be seen that the diffusion of propidium iodide inside the cells is correlated with the leak of EGFP. The results show that melittin induces the formation of pores in the plasma membranes allowing the passage of... 

Midoux, P., Kichler, A., Boutin, V., Maurizot, J.C. and Monsigny, M. (1998) Membrane permeabilization and efficient gene transfer by a peptide containing several histidines. Bioconjug. Chem., 9, 260-267. 

Midoux, P., Mayer, R. and Monsigny, M. (1995) Membrane permeabilization by alphahelical peptides a flow cytometry study. Biochim. Biophys. Acta, 1239, 249-256. 

Mechanisms of nuclear transport, 229 Membrane permeabilization, 343 Metabolic genes, 22 Metabolic instability, 216 Metallothionein, 22 Microenvironment, 443 Molossin, 350... 

A two-color dot plot (Fig. 8.23) of cells stained with propidium iodide and annexin V FITC will indicate cells in three of the four quadrants. Unstained cells are alive and well and are the double negatives they neither express phosphatidylserine on their surface nor take up propidium iodide through leaky membranes. Cells that stain just with annexin V are apoptotic they have begun to express phosphatidylserine on their surface, but have not yet gone through the process that leads to permeabilization of their cytoplasmic membrane. Cells that stain both with propidium iodide and annexin V are necrotic (that is, dead) they take up propidium iodide and also stain with annexin Y. With a permeable cell, the flow cytometer cannot tell us whether the annexin V is on the outside of the membrane (because the cells have gone through apoptosis before membrane permeabilization) or on the inside of the membrane (because the cells have died by the necrotic pathway without apoptosis). 

Physical and chemical genetic techniques have been used to enhance the permeability of yeast membranes. Permeabilizing agents, such as polymyxin B sulfate and polymyxin B nonapeptide, have been used to physically disrupt the integrity of yeast membranes (14). However, use of such chemical agents in drug screening is not ideal, because of the toxicity induced by polymyxin B treatment. 

Armah, C.N., Mackie, A.R., Roy, C., Price, K., Osbourn, A.E., Bowyer, R, Ladha, S. The membrane-permeabilizing effect of avenacinA-1 involves the reorganization of bilayer cholesterol. Biophys J 1999 76 281-290. 

FIGURE 8.10 Chronological sequence of images of a viability assay on a single Jurkat T cell, (a) Live cell perfused with trypan blue dye. Since the cell is alive, it is not stained, (b) Cell perfused with methanol, which causes cell membrane permeabilization, followed by cell death, (c) Permeabilized cell perfused with dye it is rapidly (< 5 s) stained. The entire assay is performed in less than 2 min [368]. Reprinted with permission from the American Chemical Society. 

Kuwana, T., Bouchier-Hayes, L., Chipuk, J. E., Bonzon, C., Sullivan, B. A., Green, D. R., and Newmeyer, D. D. (2005). BH3 domains ofBH3-only proteins differentially regulate Bax-mediated mitochondrial membrane permeabilization both directly and indirectly. Mol. Cell 17, 525—535. 

Following the outer mitochondrial membrane permeabilization, the apoptogenic factors are released into the cytoplasm. Among them, cytochrome c has an important role in caspase activation, because it is the cofactor for assembling a large caspase 9 activating complex in the cytoplasm, called apoptosome. Along with cytochrome c, the Apaf-1 protein and dATP or ATP are required to form this complex in the cytoplasm (Hill et al., 2003). 

Balakrishna, R., Wood, S.J., Nguyen, T.B., Miller, K.A., Suresh Kumar, E.V., Datta, A., David, S.A. Structural correlates of antibacterial and membrane-permeabilizing activities in acylpolyamines, Antimicrob. Agents Chemother 50 (2006) 852-861. 

Chipuk JE, Kuwana T, Bouchier-Hayes L, Droin NM, Newmeyer DD, Schuler M, et al. Direct activation of Bax by p53 mediates mitochondrial membrane permeabilization and apoptosis. Science 2004 303 1010-1014. 

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