Triton X-100 Molecules Research Articles

Observation of an Intact Noncovalent Homotrimer of Detergent-solubilized Rat Microsomal Glutathione Transferase-1 by Electrospray Mass Spectrometry

Microsomal glutathione transferase-1 (MGST1) is a membrane-bound enzyme involved in the detoxification of xenobiotics and the protection of cells against oxidative stress. The proposed active form of the enzyme is a noncovalently associated homotrimer that binds one substrate glutathione molecule/trimer. In this study, this complex has been directly observed by electrospray mass spectrometry analysis of active rat liver MGST1 reconstituted in a minimum amount of detergent. The measured mass of the homotrimer is 53 kDa, allowing for the mass of three MGST molecules in complex with one glutathione molecule. Collision-induced dissociation of the trimer complex resulted in the formation of monomer and homodimer ion species. Two distinct species of homodimer were observed, one unliganded and one identified as a homodimer.glutathione complex. Activation of the enzyme by N-ethylmaleimide through modification of Cys(49) (Svensson, R., Rinaldi, R., Swedmark, S., and Morgenstern, R. (2000) Biochemistry 39, 15144-15149) was monitored by the observation of an appropriate increase in mass in both the denatured monomeric and native trimeric forms of MGST1. Together, the data correspond well with the proposed functional organization of MGST1. These results also represent the first example of direct electrospray mass spectrometry analysis of a detergent-solubilized multimeric membrane protein complex in its native state.

Monoterpenes affect chlorodiazepoxide–micelle interaction through micellar dipole potential modifications

The ability of several natural terpenes to affect benzodiazepine (BZD)–micelle interaction through the membrane dipolar organization was investigated. The acid–base equilibrium of chlorodiazepoxide (CDX) and the spectroscopic behavior of the electrochromic dye merocyanine were tested in the presence and in the absence of Triton X-100 micelles (used to mimic a membrane environment) containing or not cineole, menthol, geraniol or camphor. CDX's apparent pK increased in the environment of terpene-containing micelles compared with pure Triton X-100 micelles. Decrements in electric potentials (between −111 and −128 mV with respect to pure detergent) were calculated from Boltzmann equation. This result suggested, that in the presence of terpenes, the tendency of CDXH+ to remain in the membrane phase increased. The dielectric constant (D) of the microenvironment sensed by merocyanine within Triton X-100 micelles, determined from λmax,2 of merocyanine monomer, was D=9 and increased in the presence of all the terpenes assayed (D≅11). The decrease in merocyanine partitioning (Apeak1/Apeak2 increased) also reflected an increment in the negative dipole potential. The present results suggest that terpenes contributed to the whole dipolar arrangement of the micelle with a dipole moment vector which had an intense component oriented parallel to the intrinsic dipole of the Triton X-100 molecules in the micelles. This led to a more negative environment of the interface region where CDX was located, and increased the net polarity of the deepest micelle regions sensed by merocyanine.

Catalytic Mechanism Revealed by the Crystal Structure of Undecaprenyl Pyrophosphate Synthase in Complex with Sulfate, Magnesium, and Triton

Undecaprenyl pyrophosphate synthase (UPPs) catalyzes chain elongation of farnesyl pyrophosphate (FPP) to undecaprenyl pyrophosphate (UPP) via condensation with eight isopentenyl pyrophosphates (IPP). UPPs from Escherichia coli is a dimer, and each subunit consists of 253 amino acid residues. The chain length of the product is modulated by a hydrophobic active site tunnel. In this paper, the crystal structure of E. coli UPPs was refined to 1.73 A resolution, which showed bound sulfate and magnesium ions as well as Triton X-100 molecules. The amino acid residues 72-82, which encompass an essential catalytic loop not seen in the previous apoenzyme structure (Ko, T.-P., Chen, Y. K., Robinson, H., Tsai, P. C., Gao, Y.-G., Chen, A. P.-C., Wang, A. H.-J., and Liang, P.-H. (2001) J. Biol. Chem. 276, 47474-47482), also became visible in one subunit. The sulfate ions suggest locations of the pyrophosphate groups of FPP and IPP in the active site. The Mg2+ is chelated by His-199 and Glu-213 from different subunits and possibly plays a structural rather than catalytic role. However, the metal ion is near the IPP-binding site, and double mutation of His-199 and Glu-213 to alanines showed a remarkable increase of Km value for IPP. Inside the tunnel, one Triton surrounds the top portion of the tunnel, and the other occupies the bottom part. These two Triton molecules may mimic the hydrocarbon moiety of the UPP product in the active site. Kinetic analysis indicated that a high concentration (>1%) of Triton inhibits the enzyme activity.

Preparation of silver nanoparticles in hexagonal phase formed by nonionic Triton X-100 surfactant

Silver nanoparticles were prepared by reducing silver ions in the hexagonal phase formed by Triton X-100 in aqueous solution. Triton X-100 molecules have been used to form the hexagonal phase in aqueous solution as well as to reduce the silver ions into silver atoms. The hexagonal phase hindered the growth and aggregation of particles. The microstructure of the hexagonal phase was investigated by polarizing microscopy, 2H NMR spectroscopy and small-angle X-ray diffraction. At the initial stage of the reaction, silver particles prepared in the hexagonal phase exhibited a size of 1–7 nm. As the reaction proceeded, particles grew up to about 30 nm as determined by transmission electron microscopy (TEM). The formation rate of silver particles was investigated by UV–Visible spectroscopy. It was found that the reaction temperature was an important factor for the rate of particle formation. With TEM, it was confirmed that surfactant aggregates, which have flexible structures, could not absolutely prevent particles from growing and aggregating. But, the results of polarizing microscopy, 2H NMR spectroscopy and X-ray diffraction exhibited that the growth of particles could not cause a deformation of the original structure (hexagonal phase), which was employed as the reaction medium.

Self-Assembly of Dendritic Micellar Structures Based on Triton X-100 and Branched Poly(ethylene oxide)-containing Cyclodextrins

Interaction between the branched poly(etylene oxide)-containing cyclodextrins and nonionic surfactant Triton X-100 was studied. Amphiphilic character of substituted cyclodextrins where poly(etylene oxide) fragments serve as the hydrophilic moiety and outer cyclodextrin surface, as the hydrophobic moiety. Using two groups of experimental methods, it was shown that substituted β-cyclodextrin forms inclusion complexes with hydrophilic poly(ethylene oxide) fragment of Triton X-100 molecule, surfactant molecules being incorporated into a micelle composition. The first group of these methods includes viscometry of solutions and measurement of rotational diffusion coefficient, while the other group (tensiometry, gel permeation chromatography, and spectrophotometry) makes it possible to determine colloidal properties of components. The modified micelles consist of nonionic surfactant molecules, whose outer ends are strung with molecules of substituted cyclodextrins; moreover, their structure resembles that of dendrimers due to arising branchings. Note that the self-assembly method was employed to prepare such dendrimers for the first time.

Crystal structure of the liganded SCP-2-like domain of human peroxisomal multifunctional enzyme type 2 at 1.75 Å resolution

β-Oxidation of amino acyl coenzyme A (acyl-CoA) species in mammalian peroxisomes can occur via either multifunctional enzyme type 1 (MFE-1) or type 2 (MFE-2), both of which catalyze the hydration of trans-2-enoyl-CoA and the dehydrogenation of 3-hydroxyacyl-CoA, but with opposite chiral specificity. MFE-2 has a modular organization of three domains. The function of the C-terminal domain of the mammalian MFE-2, which shows similarity with sterol carrier protein type 2 (SCP-2), is unclear. Here, the structure of the SCP-2-like domain comprising amino acid residues 618-736 of human MFE-2 (dΔhΔSCP-2L) was solved at 1.75 Å resolution in complex with Triton X-100, an analog of a lipid molecule. This is the first reported structure of an MFE-2 domain. The dΔhΔSCP-2L has an α/β-fold consisting of five β-strands and five α-helices; the overall architecture resembles the rabbit and human SCP-2 structures. However, the structure of dΔhΔSCP-2L shows a hydrophobic tunnel that traverses the protein, which is occupied by an ordered Triton X-100 molecule. The tunnel is large enough to accommodate molecules such as straight-chain and branched-chain fatty acyl-CoAs and bile acid intermediates. Large empty apolar cavities are observed near the exit of the tunnel and between the helices C and D. In addition, the C-terminal peroxisomal targeting signal is ordered in the structure and solvent-exposed, which is not the case with unliganded rabbit SCP-2, supporting the hypothesis of a ligand-assisted targeting mechanism.

Mixed Micelles of Triton X-100 and Cetyl Trimethylammonium Bromide in Aqueous Solution Studied by 1H NMR

1H NMR chemical shift, spin−lattice relaxation time, spin−spin relaxation time, and two-dimensional nuclear Overhauser enhancement (2D NOESY) measurements show that Triton X-100 (TX-100) molecules coaggregate with cetyl trimethylammonium bromide (CTAB) molecules in aqueous solutions. The concentration of TX-100 in the mixed solutions of this study is 3 mM, with varying molar ratios of CTAB/TX-100 (C/T) ranging from 0.5 to 2.9. The results give information about the structure of the mixed micelles. The α-methylene group of CTAB is in the near vicinity of the phenoxy ring of TX-100. The trimethyl group attached to the polar head of CTAB locates between the first oxyethylene group next to the phenoxy ring of TX-100, and the end methyl group of CTAB is close to those of TX-100. The closely packed (coiled) hydrophilic polyoxyethylene chains in the exterior part of the mixed micelles gradually extend with an increase in C/T in the mixed solution. CTAB and TX-100 molecules are uniformly mixed in the micelles in ...

Self-aggregation of surfactants in water solution by NMR

Dynamic behavior of surfactant molecules, sodium dodecyl sulfonate (SDSN), cetyl trimethyl ammonium bromide (CTAB) and triton X-100, and their micelles in water solution at various concentrations were studied by chemical shift, self diffusion coefficient (D) and nuclear magnetic resonance relaxation (T1 and T2) measurements. Experimental results show that there exist turning points at surfactant concentration (C) nearby the critical micelle concentration (cmc) for all of the above-mentioned parameters, which approach the constant values when C equals 5 to 10 times cmc. T1 and T2 measurements show that when C<cmc, Triton X-100 molecules exhibit fast isotropic motion in water solution. After micellization, the motion of hydrophobic chains is far away from the extreme narrowing condition, however, the long hydrophilic oxyethylene chains move relatively freely, although their T1 and T2 become shorter in comparison with those of the molecules in mono-molecular state. The behavior of the hydrophobic chains within the micellar core of SDSN and CTAB is similar to that of Triton X-100 to some extent.

Observation of the Dynamic and Collective Behavior of Surfactant Molecules at a Water/Nitrobenzene Interface by a Time-Resolved Quasi-Elastic Laser-Scattering Method

A spectroscopic technique of time-resolved quasi-elastic laser-scattering measurement was applied to monitor the dynamic and collective behavior of anionic surfactant sodium dodecyl sulfate (SDS) and neutral surfactant Triton X-100 molecules around their critical micelle concentrations (cmc's) at a water/nitrobenzene liquid/liquid interface. The time courses of the capillary-wave frequencies after the injection of the surfactant solutions into the water phase were investigated. We found that the dynamic and collective behavior of these two kinds of surfactant molecules at the interface differ from that of the cationic surfactant cetyltrimethylammonium bromide we reported previously. Our results suggested that for SDS the monomolecules at the interface coexist with some molecular aggregates such as micelles at the interface; for Triton X-100, the monolayer at the interface was disrupted and some molecular aggregates, such as micelles, were formed at the interface.

Study of spontaneous insertion of non-ionic surfactant Triton X-100 into phospholipid monolayers

L- α- Phosphatidylcholine (DOPC, Dioleoyl) monolayers of different initial surface pressures were spread at the sub-phase surface of a home-made Teflon mini-trough. Surface pressure was measured by the Wilhelmy plate method so as to study the spontaneous membrane insertion of non-ionic surfactant Triton X-100. A formula was deduced to approximately give out the final insertion amounts of Triton X-100 under different initial surface pressures. The results showed that the insertion amount decreased as the initial surface pressure increased and the amount included an `excess' of Triton X-100, which could be used to approximately give out the interaction energy quantity between lipid and Triton X-100 molecules in the monolayer.

Reverse Micelle Systems Composed of Water, Triton X-100, and Phospholipids in Organic Solvents: 1. Phase Boundary Titrations and Dynamic Light Scattering Analysis

Enzymes entrapped in systems formed with water, phospholipids, toluene, and Triton X-100 show a catalytic activity that is much lower and a thermostability that is much higher than that observed in totally aqueous systems or in other types of reverse micelles. By phase boundary titrations and dynamic light scattering, this work characterizes reverse micelle systems formed in either toluene or propylbenzene with Triton X-100 and water. Four regions with distinct structural features were encountered. Up to one molecule of water per one Triton X-100 molecule, the system was transparent; light scattering measurements of this region indicated that water hydrated Triton X-100 monomers. A turbid region was formed as water content was increased to water:Triton X-100 ratios of 7.6 in toluene and 4.2 in propylbenzene. This thermodynamically unstable region was formed by large polydisperse structures. Transparent systems containing small size (27–150 Å) thermodynamically stable reverse micelles were formed when the ratio of water to Triton X-100 molecules in the reverse micelle was in the range of 7.6 to 26.8 in toluene and 4.2 to 15.1 in propylbenzene. In this region, micellar size increased with water content. Water concentrations higher than the latter values resulted in phase separation. A similar titration of the aforementioned systems in the presence of phospholipids revealed that in the first region of transparency up to 10 molecules of water hydrated a phospholipid molecule. The inclusion of phospholipids to the Triton X-100 systems caused a displacement of the boundaries of the second region of transparency toward higher water contents.

Positronium reactions in the triton X-100-p-nitro phenol system

The positronium reactions have been used as a probe to study the solubilization of p-nitro phenol within the micellar phase of a Triton X-100 solution and the corresponding changes in the molecular association phenomenon. The presence of p-nitro phenol resulted in an enhancement of micellization which has been corroborated by surface tension measurements. This paper also lays emphasis on the secondary aggregation phenomenon of Triton X-100 molecules at the far post-micellar stage (≈10–15 mM). The solubilization of p-nitro phenol at various stages of aggregation has been discussed through the interaction with positronium atoms by setting up a kinetic model and reaction equilibria.

The Adsorption of Triton X-100 at the Air-Aqueous Solution Interface

Measurements of the surface tension of aqueous solutions of Triton X-100 (TX-100) were made at different temperatures. Isotherms and thermodynamic adsorption parameters were determined from the surface tensions. Also, the equation of state for the monolayer film of TX-100 at the water-air interface was analyzed. A linear dependence was found between log Γ and log C in the region of low concentration of TX-100 in solution, and three different methods used to calculate the free energy of adsorption gave slightly different results. In the region of low concentration the Π(A - A 0 ) values were equal to 1kT. At high concentration, there were attractive Lifshitz-van der Waals forces between the apolar parts of the TX-100 molecules and repulsive acid-base forces between the heads of the TX-100 molecules in the adsorbed monolayer film.

Adsorption kinetics of octylphenyl ethers of poly(ethylene glycol)s on the solution—air interface

We studied the dynamic surface tension of aqueous solutions of Triton X-100 and Triton X-405 by the maximum bubble pressure and the inclined plate methods in the lifetime range from 0.001 s up to 10 s. It is established that in the region of large and ultimately small surface pressure and time the adsorption follows diffusion kinetics, but in the region of intermediate values of lifetime and surface pressure both the surfactants decrease the surface tension faster than predicted by the existing diffusion theory. We offer a model that provides for the ability of poly(ethylene glycol) chains to adsorb on the water-air interface and to change the area that a molecule occupies on the surface. For this model we achieve full coincidence of the measured values and the values calculated according to the diffusion theory of the dynamic surface tension. It is ascertained that the Triton X-405 molecule can exist in the surface layer in different states: with the poly(ethylene glycol) chain fully expanded or with it partially or fully submerged in solution. The first state is most probable at surface pressures less than 5 mN m −1, and the second is probable at a pressure of about 8–10 mN m −1. At pressures larger than 15–20 mN m −1, the poly(ethylene glycol) chain is fully submerged in the solution. The Triton X-100 molecule can also expand its poly(ethylene glycol) chain at low pressures and fully submerges it in the solution at higher values of the surface pressure.

Cyclic voltammetric study of mixed monolayers of methylene blue and Triton X-100 formed spontaneously on mercury

We studied the spontaneous formation of mixed monolayers of methylene blue (MB +) and Triton X-100 (TX) from aqueous solutions containing both substances and a fixed concentration of MB + of c MB = 2.5 × 10 −6 M. The voltammetric peak obtained under these conditions was typical of redox sites immobilized at the electrode and was ascribed to the reduction of MB + molecules dissolved in the aliphatic subphase of the surface film formed by the TX molecules. This phenomenon arises from the hydrophobic character of MB + molecules and gives rise to strong interactions between such molecules and the hydrophobic moiety of TX molecules. The voltammetric peak of MB + in the surface film is of a quasi-reversible nature. The transfer coefficients of the reduction and oxidation processes were found to be aα f n ⋍ 1 and α b n ⋍ 1.5, with n = 2. A plausible mechanism to account for the relationship α f n + α b n ≠ 2 is put forward.

Effects of reconstitution of membrane-bound lipoprotein lipase and dispersity of substrate on its reaction characteristics

Reaction characteristics of a membrane-bound lipoprotein lipase acting on a hydrophobic substrate were investigated in aggregated structures—lipid bilayers of liposomes and mixed micelles of Triton X-100. The enzyme activity was enhanced with increases in Triton X-100 and phospholipid concentrations in micellar and liposomal structures. This higher activity was found to be due to both the solubilization state of the hydrophobic substrate and the hydrophobic interactions of the enzyme with either phospholipid or Triton X-100 molecules as a result of its incorporation into the aggregated systems. The enzyme reconstituted into lipid bilayers of liposomes prepared from 15 mM DMPC in the presence of 0.05% Triton X-100 showed a further 1.5-fold higher activity in comparison with the activity without reconstitution in micelles of 1.0% Triton X-100. These results indicate the necessity of the bilayer structure to retain the membrane-bound enzyme in an active conformation.

Multinuclear NMR characterization of CTAB—hexanol—water, sodium oleate—butanol—water and triton X-100—decanol—water microemulsions

Multinuclear NMR is a very valuable tool to characterize micellar systems or microemulsions. It allows one to determine c.m.c. values, to study the dissolution of organic molecules, the salvation of cations and anions, the structural changes occurring in a ternary diagram, the mobility of the molecules, etc. This review paper essentially deals with the characterization of cationic (CTAB—hexanol—water), anionic (sodium oleate—butanol—water) and neutral (Triton X-100—decanol—water) reversed micelles. The use of paramagnetic ions [Ni(II), Co(II), Fe(III), etc.] is particularly emphasized to characterize the site of solubilization and their interaction with surfactant and cosurfactant molecules ( 13C-NMR). It is concluded, that the metallic ions are basically solvated in the inner water cores and one or more hexanol molecules are included in their first coordination shells in the CTAB—hexanol—water microemulsions. In the Triton X-100—decanol—water microemulsions, both decanol and Triton X-100 molecules enter the first coordination shell of Co(II) ions which are dissolved in both aqueous water cores and the organic medium. 19F-NMR of a fluorinated probe molecule is particularly useful to study the size of the inner water cores. The method is based on the partition of the molecules between the interface and the organic medium. However, this method has to be applied with great care, and the computed data have to be compared to other physico-chemical results. Both 19F-and 23Na-NMR results show a great variation of the behaviour of the sodium oleate—butanol—water system in the so-called bicontinuous region. The Na + ions are reoriented independently on a hypothetical inverse micellar droplet.

Reconstitution of isolated Rieske Fe-S protein into a Rieske-depleted cytochrome b6-ƒ complex

The Rieske Fe-S protein can be isolated from the cytochrome b 6-ƒ complex by means of chromatography on a hydroxyapatite column in the presence of detergent. Depletion of the cytochrome complex from the Rieske protein results in the loss of oxidoreductase activity, as well as the ability to reduce cytochrome b 6. The Rieske Fe-S protein can be reconstituted into the Rieske-depleted complex by removal of the Triton X-100 molecules associated with the protein fractions, and their substitution by lipids. Upon reconstitution the complex is reactivated, and the role of the Rieske Fe-S protein in the reduction of both plastocyanin and cytochrome b 6 can be demonstrated.

Detergents inhibit exocytosis in PC 12 cells: evidence for an effect on ion fluxes.

Membrane events in exocytosis were studied by examining the effect of different detergents on the K+-stimulated release of noradrenaline in the secretory cell line PC 12. The nonionic detergent Triton X-100 and the cationic detergent cetyltrimethylammonium bromide (CTAB) inhibit the noradrenaline release evoked by 55 mM K+ by 50% at very low concentrations (30 microM and 10 microM, respectively). These values are tenfold lower than the critical micellar concentrations (CMC). No such effect was seen with the anionic detergent sodium dodecyl sulphate (NaDodSO4). The inhibitory effect of 30 microM Triton X-100 is reversible, and the recovery from inhibition correlates with the loss of detergent from the cells as demonstrated by binding studies using [3H]Triton X-100. The possible relationship between this inhibition of secretion and the structural properties of the detergent was investigated. The inhibition in the presence of purified Triton X-100 subfractions turned out to be a function of the length of the oligometric ethyleneglycol chain (C6 to C26). The maximal effect was observed for Triton X-100 molecules having a chain length of 16 carbon atoms, which can penetrate just half of the lipid bilayer of the membrane. Additionally, the phase transition at 13-14 degrees C observed in an Arrhenius plot of noradrenaline release in stimulated cells was abolished. In the presence of 30 microM Triton X-100, 22Na+ uptake, 86Rb+ release, and 45Ca2+ uptake were reduced by 50-60%. These data suggest that the site of action of Triton X-100 is at the level of altering the movement of ions in PC 12 cells during the stimulatory phase of secretion.

ChemInform Abstract: Raman Spectra and Molecular Conformation of 2,4,4‐Trimethyl‐2‐pentanethiol as a Model Compound of a Hydrophobic Group of Triton X‐100 Surfactant.

Abstract Raman spectra of 2,4,4-trimethyl-2-pentanethiol were measured. The spectral analysis with the normal coordinate treatment indicated that this molecule takes the gauche conformation about the CCCS bond in the solid state and the trans and gauche conformations in the liquid state. The Raman bands due to the totally symmetric C&.zdbnd;C streching vibration of the t-butyl part of the 1,1,3,3-tetramethylbutyl group were found to be important to distinguish the two conformations. These key bands were applied to the interpretation of the Raman spectra of Triton X-100 surfactant which contains the p -(1,1,3,3-tetramethylbutyl)phenoxyl group as a hydrophobic moiety. The 1,1,3,3-tetramethylbutyl group of Triton X-100 molecules is shown to be predominantly in the gauche conformation in the liquid state and in aquaeous solution.