Triton membrane disruption

To check if PemB is surface exposed, E. chrysanthemi cells were subjected to proteolysis. Treatment of the cell suspension with trypsin, proteinase K or chimotrypsin at a concentration of 0.1 to 1 mg/ml for 1 h did not cause PemB proteolysis or its liberation into the medium. Cell pre-treatment with EDTA-lysozyme, which renders the periplasmic proteins accessible to proteases, gave no effect. PemB was also resistant to proteolytic digestion in extract of cells disrupted by sonication or in a French press. Only addition of Triton X-100 (up to 0.1%) causing formation of the micelles with PemB lead to a quick proteolyis of this protein (data not shown). In another approach to analyse the PemB exposition, bacterial cells were labelled with sulfo-NHS-biotin. This compound is unable to cross membranes and biotinylation... 

The addition of zwitterionic or non-ionic detergents to SD buffers has three important functions. Detergents aim to disrupt membranes and consequently solubilize lipids and delipidate proteins, which are bound to membranes or vesicles. The use of SDS as a detergent will affect the overall net charge and so non-ionic or zwitterionic detergents such as Triton X-100 or CHAPS are used (Shaw and Riederer, 2003). Commonly used CHAPS, which will not affect the overall charge, can be added up to 4% in SD buffers to account for... 

Since the 28K-proacrosin interaction required either 8 M urea or 6 M guanidine HC1 to disrupt it (10), this protein-protein interaction may act to anchor proacrosin to the acrosomal membranes or to other proacrosin molecules. While acrosin does not fit all the criteria for an intrinsic membrane protein, acrosin was associated with and extracted from preparations of sperm membrane vesicles with 1 mM HC1 and Triton X-100 (5 , Urch, unpublished observations). Salt or combinations of salt and detergent were ineffective. The interaction of acrosin with membranes, particularly sperm acrosomal membranes, is obviously important in ZP binding and penetration. 

We have speculated on but do not understand the mechanism causing the lytic activity of laetisaric acid. The active twelve carbon metabolite of laetisaric acid may poison a key enzyme in lipid metabolism or disrupt the integrity of the fungal cell membrane by insertion or dissolution as has been shown in Escherichia coli with sodium dodecyl sulfate and Triton X-100 (24 r 25). Why the C-12 molecule is most active remains to be determined. Kinetic studies of lipid metabolism and physicochemical and ultrastructural investigations of membranes treated with the putative active metabolite may answer these questions. 

If ChAc was released from disrupted synaptosomes by adjustment of ionic strength and pH, it could be re-bound to membranes by passing the suspension through Sephadex columns, thereby altering the ionic environments (Fig. 2). Treatment of the re-bound enzyme with Triton X-100 showed that the enzyme was present in a non-... 

The second question depends on whether AChE is present on the inside or the outside of the nerve terminals. Most people working with subcellular fractionation have obtained evidence to show that the main part of the AChE must be on the outside (Fonnum, Rodriguez de Lores Amaiz and March-banks, independent studies, unpublished observations). This is mainly based on two kinds of study. (1) If the cholinesterase activity of isolated synaptosomes is measured in sucrose, in water (hypotonic disruption) or by treatment with a detergent such as Triton X-100, the enzyme activity remains unaltered, i.e., the enzyme, unlike ChAc, is not protected behind the membrane. (2) If thesynaptosome is loaded with labelled ACh, the latter is not hydrolysed but if the synaptosomes are broken, ACh diffuses out and is immediately hydrolysed by the AChE. If there is any AChE on the inside of the nerve terminal it must therefore be only a very small amount. 

It is of interest not only to perforate vesicle membranes but also to destroy them after they have served their purpose as transport vehicles, in particular for DNA. Natural vesicles, so-called endosomes, contain about 50% cholesterol. The disruption of such cholesterol-containing lipid bilayers by Triton XI00 or sodium deoxycholate, examples of artificial and natural detergents, results in a leaky membrane at low concentration and in a catastrophic rupture process above the cmc of the amphiphiles. Vesicles made of fluid phospholipid bilayers devoid of cholesterol showed only leakiness under the same conditions. Amphiphiles with a carboxylate end group and a very bulky hydrophobic end (e.g., with two tert. butyl groups) disrupt membranes at pH 5 and have no effect above pH 7 (harpoons). For an example, see Figure 6.5.3. 

Many drug carriers are made of hydrophobic materials such as lipids and poly(butyl cyanoacrylate). It will be thermodynamically unstable for submicron particles made of these materials to remain dispersed in an aqueous environment such as blood circulation. Surfactants or block co-polymers are therefore routinely included in these formulations to prevent particle aggregation. Studies showed that a number of these agents, most noticeably the nonionic surfactants such as polysorbates (also known as Tweens) and Tritons and block co-polymers such as poloxamers (also known as Pluronics), may inhibit the ABC transporters [97-99]. As previously discussed, ABC transporters interact with their substrates in the lipid bilayers of the plasma membrane. Surfactants can disrupt the arrangement of the lipid bilayer expressing the transporters and subsequently inhibit their drug efflux activities [97, 100]. It... 

Triton X-100 and X-114 are industrial p-rcrr-octylphenol polyoxyethylene surfactants with about 9.2 and 7.5 ethylene oxide groups per molecule respectively. They are used in biochemical studies because of their ability to disrupt biological membranes without denaturing integral membrane proteins [ 119]. TX-100 [ 120] forms an H1 phase between 37 and 63% surfactant from 0 to... 

Changes in plastid membrane permeability during development also have been noted [13]. General, there is more rapid uptake of amino adds early followed by a decline as tissues mature this in turn could influence p C]aspartate incorporation rates seen in intact plastids. Again, this appears unlikely, since amino add production by Triton X-100 disrupted plastids (which would not be subject to permeability effects) varies in a similar fashion to light-driven activity (Fig. 2). 

Cells can also be disrupted chemically by detergents, osmotic shocks, organic solvents, and alkali treatments (Prasad, 2010). Detergents are molecules with hydrophilic and hydrophobic properties that allow them to interact both with water and lipids. The hydrophobic fnnction is nsnally ionic, whereas the hydrophobic aspect is normally a hydrocarbon (Joesten et al., 2006 Marriott and Gravani, 2006 Tadros, 2005). They disrupt cell membranes by penetrating between the layers and forming micelles that separate lipids and proteins. Many detergents are available for cell membrane solubilization such as sodium dodecyl sulfate and triton X. However, many of them, such as sodium dodecyl sulfate, break protein-protein interaction and denature the enzyme. 

Viewing the fact that only a portion (but not all of the acid phosphatase) of the lysosomal fraction is readily released upon physical disruption of the lysosomal membrane by freezing and thawing or by hypoos-motic pressure, Baccino et al. (1971) suggested that at least two varieties of acid phosphatase were associated with the lysosomal fraction, the first being readily, and the other not readily, dissociable from lysosomal structures. This interpretation coincides with the earlier observation of Sloat and Allen (1969) who showed two varieties of acid phosphatase associated with lysosomal fractions of rat liver. One form is readily released after physical disruption of lysosomal fractions, the other form is associated with the lysosomal membrane and became soluble only with 5% Triton X—100 treatment. This membrane-associated enzyme accounted for 40% of the total lysosomal acid phosphatase, is heat-stable, and can be separated from the soluble form by electrophoresis. But these two enzymes have similar pH optima and a common response to inhibitions by L-tartrate, fluoride, alloxan, and formaldehyde. 

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