Lauryldimethylamine Oxide Research Articles

Stabilization and strengthening effects of filamentous nanocellulose in the foam forming of quartz paper

Compared with traditional papermaking, foam forming is a new papermaking technology that uses foam instead of water to disperse fibres, which can effectively solve the problem of poor evenness of ceramic paper, but the instability of foam itself affects the application of foam forming technology. Herein, a highly stable foaming agent for foam forming technology was prepared via physical reaction of lauryl dimethyl amine oxide (OB-2) with filamentous nanocellulose (cellulose nanofiber (CNF-C) and bacterial cellulose (BC)). Then, the quartz paper was prepared by foam forming technology. Firstly, hydrogen bond interactions between hydroxyl groups of the filamentous nanocellulose and hydrophilic moieties on OB-2 enabled the formation of a 3D nanonetwork layer on the surface of the bubble, which extended the half-life of the bubble and effectively prevented the bubble from bursting or coalescing. Then, the foam was extruded and cracked, and the filamentous nanocellulose was retained on the quartz fibres to prepare filamentous nanocellulose/quartz fibre paper by foam forming technology. The quartz paper exhibited excellent evenness and mechanical properties. In conclusion, the research of foam forming technology is of great significance to the application and development of special paper.

Oil Detachment Mechanism in Natural Surfactant Flooding from Silica Surface: Molecular Dynamics Simulation

Summary International regulations have compelled Europe and the United States to phase out certain traditional surfactants to mitigate the use of toxic and nonbiodegradable chemicals. Sodium cocoyl propionate (SCA), as a natural surfactant with high performance, has been proved to have the potential to replace traditional surfactants in previous studies. However, its performance has not fully met practical application requirements. Therefore, in this paper, molecular dynamics (MD) simulation was used to study the detachment behavior and mechanism of SCA, lauryl dimethylamine oxide (OA-12), emulsifier OP-10, and SOO (combination of SCA, OA-12, and OP-10) on crude oil (dodecane, C12) at different temperatures (80–120°C) and salinities (20 000–200 000 mg/L). The complex interaction mechanism between surfactant molecules and C12 molecules was revealed by analyzing the simulated snapshot, radial distribution function (RDF), mean square displacement (MSD), and interaction energy. The simulated snapshot captures the conformational evolution of surfactant molecules at different time points, emphasizing the spatiotemporal and spatial changes of their dynamic behavior. A comparison of two desorption modes reveals that dispersive adsorption displacement and concentrated adsorption displacement are two possible desorption mechanisms. RDF analysis shows that the probability of SOO molecules near C12 remains high even at high-salinity and -temperature conditions. MSD analysis showed that the diffusion capacity of SOO was the highest at 100°C, reaching 1.52867×10 –5 cm2/ps. The calculation of interaction energy results reveals that SOO has a strong adsorption capacity for C12, which is mainly due to the effect of van der Waals (vdW) force. This is because the C12 molecules are inert, and their molecular movement is mainly determined by the polar groups of the surfactant. The main contribution of this study is to provide a natural surfactant with superior performance as a viable alternative, offering experimental settings for further improvement in SCA performance. This research provides theoretical guidance for on-site applications of SCA and SOO to enhance oil recovery.

Nonionic and anionic surfactants as flowback aids in hydraulic fracturing methods of crude oil production

AbstractSurfactants find utility in hydraulic fracturing operations for their ability to modify rock wettability and increase the flowback of fracturing fluid following proppant delivery. The performance of a series of nonionic and anionic surfactants was evaluated by gravity drainage displacement testing. Alkylbenzenesulfonates having alkyl chains possessing at least 16 carbons were among the best performers, yet a similar surfactant having an alkyl chain possessing only ~12 carbons performed quite poorly. These data illustrate that a critical chain length exists for this series. Lauryldimethylamine oxide also performed well. In general, those surfactants possessing the greatest ionic character outperformed nonionic surfactants. Critical micelle concentration, surface tension, contact angle, relative solubility number, and hydrophilic–lipophilic balance were determined for each surfactant studied. Unexpectedly however, few correlations between any of these physical and surface properties and performance in gravity drainage displacement tests were identified, underscoring the complexity of selecting surfactants suitable in the application. These data suggest that more real‐world test methods, employing stationary phases, temperatures and pressures that better mimic the fields and individual wells being considered, are needed to better guide surfactant selection.

Влияние различных типов поверхностно-активных веществ на эмиссию газов и микробиоту жидкой фракции навозных стоков

Various surfactants are widely used in modern industrial animal husbandry. As part of detergents and pharmacological veterinary preparations, surfactants inevitably fall into manure effluents (ME), resulting in a change in the properties of ME. The study of the effect of various types of surfactants on the gas release and microbiota of the liquid fraction (LF) of ME was carried out on the example of surfactants with different biocidal activity: sodium laureth sulfate – anionactive surfactant (AS), lauryldimethylamine oxide – nonionic surfactant (NS) and benzalkonium chloride – cationic surfactant (CS). The studies were conducted under laboratory conditions. The total observation time since the introduction of additives in the LF (0,01 g per 1 liter) was 14 days. The obtained data shows changes in the dynamics of greenhouse gases (CH4 and CO2) and odor-forming substances (H2S, NH3, RSH) emission, as well as in the composition of the microbiota of biofilms and bacterioplankton, under the influence of surfactants. The dynamics of the CH4 emission differ from those of other gases. The use of CS leads to a gradual increase in CH4 emission a week after application. This may be due to increased permeability of the cytoplasmic membranes of methanogenic archaea with increased chemical resistance. The application of various types of surfactants into the LF has different effects on the dynamics of the odor-forming substances emission, and the active emission period of mercaptans becomes longer. The highest level of ethyl mercaptan was observed in the option with NS. There was no significant difference in the effect of low concentrations of various surfactants on the microbiota of LF, however the AS has a more pronounced bacteriostatic effect on set of indicators. The most tolerant to the effects of all types of surfactants were Clostridia, which dominates the microbiota biofilms and bacterioplankton. Fungi were the second most abundant in biofilms of the LF.

Cryo-EM structure of the Mycobacterium abscessus F1-ATPase

The cases of lung disease caused by non-tuberculous mycobacterium Mycobacterium abscessus (Mab) are increasing and not reliably curable. Repurposing of anti-tuberculosis inhibitors brought the oxidative phosphorylation pathway with its final product ATP, formed by the essential F1FO-ATP synthase (subunits α3:β3:γ:δ:ε:a:b:b′:c9), into focus as an attractive inhibitor target against Mab. Because of the pharmacological attractiveness of this enzyme, we generated and purified a recombinant and enzymatically active Mab F1-ATPase complex, including subunits α3:β3:γ:δ:ε (MabF1-αβγδε) to achieve mechanistic, regulatory, and structural insights. The high purity of the complex enabled the first cryo-electron microscopy structure determination of the Mab F1-ATPase complex to 7.3 Å resolution. The enzyme showed low ATP hydrolysis activity, which was stimulated by trypsin treatment. No effect was observed in the presence of the detergent lauryldimethylamine oxide.

A many-body dissipative particle dynamics parametrisation scheme to study behaviour at air-water interfaces.

In this article, we present a general parametrisation scheme for many-body dissipative particle dynamics (MDPD). The scheme is based on matching model components to experimental surface tensions and chemical potentials. This allows us to obtain the correct surface and mixing behaviours of complex, multicomponent systems. The methodology is tested by modelling the behaviour of nonionic polyoxyethylene alkyl ether surfactants at an air/water interface. In particular, the influence of the number of ethylene oxide units in the surfactant head group is investigated. We find good agreement with many experimentally obtained parameters, such as minimum surface area per molecule; and a decrease in the surface tension with increasing surfactant surface density. Moreover, we observe an orientational transition, from surfactants lying directly on the water surface at low surface coverage, to surfactants lying parallel or tilted with respect to the surface normal at high surface coverage. The parametrisation scheme is also extended to cover the zwitterionic surfactant lauryldimethylamine oxide (LDAO), where we provide good predictions for the surface tension at maximum surface coverage. Here, if we exceed this coverage, we are able to demonstrate the spontaneous production of micelles from the surface surfactant layer.

Investigating the Lipid Selectivity of Membrane Proteins in Heterogeneous Nanodiscs.

The structure and function of membrane proteins can be significantly impacted by the surrounding lipid environment, but membrane protein-lipid interactions in lipid bilayers are often difficult to study due to their transient and polydisperse nature. Here, we used two native mass spectrometry (MS) approaches to investigate how the Escherichia coli ammonium transporter trimer (AmtB) and aquaporin Z (AqpZ) selectively remodel their local lipid environment in heterogeneous lipoprotein nanodiscs. First, we used gas-phase ejection to isolate the membrane protein with bound lipids from heterogeneous nanodiscs with different combinations of lipids. Second, we used solution-phase detergent extraction as an orthogonal approach to study membrane protein remodeling of lipids in the nanodisc with native MS. Our results showed that Triton X-100 and lauryldimethylamine oxide retain lipid selectivity that agrees with gas-phase ejection, but C8E4 distorts some preferential lipid interactions. Both approaches reveal that AmtB has a few selective binding sites for phosphatidylcholine (PC) lipids, is selective for binding phosphatidylglycerols (PG) overall, and is nonselective for phosphatidylethanolamines (PE). In contrast, AqpZ prefers either PC or PG over PE and prefers PC over PG. Overall, these experiments provide a picture of how membrane proteins bind different lipid head groups in the context of mixed lipid bilayers.

Effect of detergents on froth stability and flotation separation

Abstract Froth stability is an important factor in determining flotation yield, and one of the factors affecting froth stability is dependent on the concentration and type of surfactant. Since detergents contain different types of surfactants, it is possible that they affect the froth stability (together with the frothing agent and the collector) and consequently the flotation yield. Therefore, the effect of detergents containing lauryldimethylamine oxide and sodium laureth sulphate was investigated in this study. The results showed that the addition of detergents increased the froth stability by more than 30% and 45% in terms of froth formation and froth decay, respectively. The flotation tests also showed that the addition of detergents increased the sulphur content of the concentrate by about 0.7%.

Regulation of ATP hydrolysis by the ε subunit, ζ subunit and Mg-ADP in the ATP synthase of Paracoccus denitrificans

F1FO-ATP synthase is a crucial metabolic enzyme that uses the proton motive force from respiration to regenerate ATP. For maximum thermodynamic efficiency ATP synthesis should be fully reversible, but the enzyme from Paracoccus denitrificans catalyzes ATP hydrolysis at far lower rates than it catalyzes ATP synthesis, an effect often attributed to its unique ζ subunit. Recently, we showed that deleting ζ increases hydrolysis only marginally, indicating that other common inhibitory mechanisms such as inhibition by the C-terminal domain of the ε subunit (ε-CTD) or Mg-ADP may be more important. Here, we created mutants lacking the ε-CTD, and double mutants lacking both the ε-CTD and ζ subunit. No substantial activation of ATP hydrolysis was observed in any of these strains. Instead, hydrolysis in even the double mutant strains could only be activated by oxyanions, the detergent lauryldimethylamine oxide, or a proton motive force, which are all considered to release Mg-ADP inhibition. Our results establish that P. denitrificans ATP synthase is regulated by a combination of the ε and ζ subunits and Mg-ADP inhibition.

Bacterial Endotoxin Testing-Fast Endotoxin Masking Kinetics in the Presence of Lauryldimethylamine Oxide.

For release of parenteral drug products, bacterial endotoxin testing is one of a panel of necessary tests. In order to ensure the validity of such tests, various controls are performed, including demonstration of compendial method suitability or method qualification. In addition to compendial suitability testing, quality control (QC) sample hold-time studies are requested by authorities like the Food and Drug Administration (FDA) as described in “Guidance for Industry: Pyrogen and Endotoxins Testing.” It is requested to be determine whether the ability to detect endotoxins can be affected by storage and handling of the sample to be tested. To accomplish these studies, endotoxin is introduced or spiked into the undiluted product and held for a certain period of time in process-representative containers. This time period reflects procedural maximum QC sample hold time from sampling until analysis. Inadequate detection of endotoxin can be caused by adsorption of endotoxin to container surfaces or molecular masking effects, in which the binding sites on the endotoxin molecules are prevented from triggering the enzymatic cascade necessary in the assay, are obscured. The endotoxin may form macromolecular structures, such as sheets or blebs, or the binding sites may otherwise be rendered unavailable due to the sample matrix composition. In either case, the endotoxin assay may yield falsely low results if and when masking occurs. In this work, the QC sample hold times of different in-process controls within the production process of a biopharmaceutical product were analyzed. One out of eight different samples showed a strong masking of endotoxin. Analysis of the sample composition revealed that either kifunensine, mycophenolic acid (MPA), or lauryl-N, N-dimethylamine oxide (LDAO) was responsible for masking. Further analysis clearly identified LDAO as the root cause for masking. A novel one-step mechanism for LDAO-induced endotoxin masking is proposed. The principle is similar to an already-proposed two-step mechanism for endotoxin masking, but the LDAO case combines these two steps: the disturbance of the salt bridges and hydrophobic interactions with LPS in one molecule. These molecular interactions occur quickly when both endotoxin and LDAO are present in the same matrix. Thus, depending on the masking agents, low endotoxin recovery (LER) can occur regardless of the QC sample hold duration.

Intensity Ratio of LH2 Complexes from Rhodopseudomonas palustris and Rhodobacter sphaeroides for Purity Determination

Purification procedures to obtain light-harvesting complexes were performed on two photosynthetic bacteria, i.e. Rhodospseudomonas (Rps.) palustris and Rhodobacter (Rb.) sphaeroides. In this study, purification was initially carried out using series of centrifugation towards the bacteria cell to acquire chromatophore of the bacteria. Application of detergent (lauryldimethylamine oxide, or LDAO) to the chromatophore enabled the extraction of membrane containing photosynthetic apparatus. Further purification involved the application of sucrose density gradient centrifugation and ion exchange chromatography. The purity of collected LH2 complexes can be determined by calculating the ratio of AB850 : Aprotein. The resulting LH2 complexes from both bacteria showed relatively high ratio suggesting the purity of the complexes.

Surfactant Effects on the Permeability of Photosynthetic Membrane from Rhodobacter sphaeroides 2.4.1 Probed by Electrochromic Shift of Endogenous Carotenoids

Four surfactants, sodium cholate(SC), n-dodecyl-β-D-maltopyranoside(DDM), lauryldimethylamine oxide(LDAO) and Triton X-100(TX), which are generally used in photosynthetic pigment-protein complexes preparation, were studied on their interaction with photosynthetic membrane from Rhodobacter sphaeroides 2.4.1 by electrochromic absorption band-shift of endogenous carotenoids and by vesicle size measurements as well. The surfactant critical micelle concentration(cmc) was found to be negatively correlated with the capability of enhancing the permeability of photosynthetic membranes to proton, and more elaborated model of surfactants interacting with membranes was obtained. The electrochromic absorption band-shift measurement might develop into a useful tool to evaluate the effects of surfactants on various membranes.

Development and validation of an analytical procedure for quantitation of surfactants in dishwashing detergents using ultra-performance liquid chromatography-mass spectrometry

The detection of surfactants in dishwashing detergents is challenging because the detergents tend to create a lot of foam. The aim of this study was therefore to develop a simple and robust procedure for simultaneous identification and quantification of multiple surfactants in dishwashingdetergents using standard addition method and ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). The results contain the method validation for 11 surfactants (linear alkylbenzene sulfonates, PPG-9-ethylhexeth-5, sodium lauryl polyoxyethylene ether sulfate, alkyl polyglucoside, sodium C14–16 olefin sulfonate, cocamide diethanolamine, cocamide monoethanolamine, lauryldimethylamine oxide, cocamidopropyl betaine, nonylphenol, and nonylphenol ethoxylate). The correlation coefficients of the linear calibration curves were larger than 0.98. The limits of detection and quantification were in the ranges of 0.43–8.0 μg/mL and 1.42–26.4 μg/mL, respectively. The precision ranged from 1.72% to 11.70%. The recoveries were within the range of 84–115%. The validation criteria were fulfilled for the 11 tested surfactants. The analytical procedure was applied to the analysis of 20 dishwashing detergent products collected from markets. The surfactants labeled on 13 out of the 20 products were consistent with the test results. Two surfactants, nonylphenol and nonylphenol ethoxylate, are relatively toxic to aquatic plants, fish, and aquatic invertebrates. These two surfactants were not detected among the 20 dishwashing detergent products, indicating that they were likely not added to the dishwashing detergent products. The developed UPLC-MS-based analytical procedure can be effectively applied to surfactant identification and quantification in dishwashing detergent products.

ADP-Inhibition of H+-FOF1-ATP Synthase.

H+-FOF1-ATP synthase (F-ATPase, F-type ATPase, FOF1 complex) catalyzes ATP synthesis from ADP and inorganic phosphate in eubacteria, mitochondria, chloroplasts, and some archaea. ATP synthesis is powered by the transmembrane proton transport driven by the proton motive force (PMF) generated by the respiratory or photosynthetic electron transport chains. When the PMF is decreased or absent, ATP synthase catalyzes the reverse reaction, working as an ATP-dependent proton pump. The ATPase activity of the enzyme is regulated by several mechanisms, of which the most conserved is the non-competitive inhibition by the MgADP complex (ADP-inhibition). When ADP binds to the catalytic site without phosphate, the enzyme may undergo conformational changes that lock bound ADP, resulting in enzyme inactivation. PMF can induce release of inhibitory ADP and reactivate ATP synthase; the threshold PMF value required for enzyme reactivation might exceed the PMF for ATP synthesis. Moreover, membrane energization increases the catalytic site affinity to phosphate, thereby reducing the probability of ADP binding without phosphate and preventing enzyme transition to the ADP-inhibited state. Besides phosphate, oxyanions (e.g., sulfite and bicarbonate), alcohols, lauryldimethylamine oxide, and a number of other detergents can weaken ADP-inhibition and increase ATPase activity of the enzyme. In this paper, we review the data on ADP-inhibition of ATP synthases from different organisms and discuss the in vivo role of this phenomenon and its relationship with other regulatory mechanisms, such as ATPase activity inhibition by subunit ε and nucleotide binding in the noncatalytic sites of the enzyme. It should be noted that in Escherichia coli enzyme, ADP-inhibition is relatively weak and rather enhanced than prevented by phosphate.

β-グリチルレチン酸／ラウリルジメチルアミンオキシド水溶液のミセル構造解析

β-グリチルレチン酸／ラウリルジメチルアミンオキシド水溶液のミセル構造解析

Biophysical Characterization of a Thermoalkaliphilic Molecular Motor with a High Stepping Torque Gives Insight into Evolutionary ATP Synthase Adaptation

F1F0 ATP synthases are bidirectional molecular motors that translocate protons across the cell membrane by either synthesizing or hydrolyzing ATP. Alkaliphile ATP synthases are highly adapted, performing oxidative phosphorylation at high pH against an inverted pH gradient (acidin/alkalineout). Unlike mesophilic ATP synthases, alkaliphilic enzymes have tightly regulated ATP hydrolysis activity, which can be relieved in the presence of lauryldimethylamine oxide. Here, we characterized the rotary dynamics of the Caldalkalibacillus thermarum TA2.A1 F1 ATPase (TA2F1) with two forms of single molecule analysis, a magnetic bead duplex and a gold nanoparticle. TA2F1 rotated in a counterclockwise direction in both systems, adhering to Michaelis-Menten kinetics with a maximum rotation rate (Vmax) of 112.4 revolutions/s. TA2F1 displayed 120° unitary steps coupled with ATP hydrolysis. Torque measurements revealed the highest torque (52.4 piconewtons) derived from an F1 molecule using fluctuation theorem. The implications of high torque in terms of extreme environment adaptation are discussed.

Deuterated detergents for structural and functional studies of membrane proteins: Properties, chemical synthesis and applications.

Detergents are amphiphilic compounds that have crucial roles in the extraction, purification and stabilization of integral membrane proteins and in experimental studies of their structure and function. One technique that is highly dependent on detergents for solubilization of membrane proteins is solution-state NMR spectroscopy, where detergent micelles often serve as the best membrane mimetic for achieving particle sizes that tumble fast enough to produce high-resolution and high-sensitivity spectra, although not necessarily the best mimetic for a biomembrane. For achieving the best quality NMR spectra, detergents with partial or complete deuteration can be used, which eliminate interfering proton signals coming from the detergent itself and also eliminate potential proton relaxation pathways and strong dipole-dipole interactions that contribute line broadening effects. Deuterated detergents have also been used to solubilize membrane proteins for other experimental techniques including small angle neutron scattering and single-crystal neutron diffraction and for studying membrane proteins immobilized on gold electrodes. This is a review of the properties, chemical synthesis and applications of detergents that are currently commercially available and/or that have been synthesized with partial or complete deuteration. Specifically, the detergents are sodium dodecyl sulphate (SDS), lauryldimethylamine-oxide (LDAO), n-octyl-β-D-glucoside (β-OG), n-dodecyl-β-D-maltoside (DDM) and fos-cholines including dodecylphosphocholine (DPC). The review also considers effects of deuteration, detergent screening and guidelines for detergent selection. Although deuterated detergents are relatively expensive and not always commercially available due to challenges associated with their chemical synthesis, they will continue to play important roles in structural and functional studies of membrane proteins, especially using solution-state NMR.

Is the incidence of aliphatic amine-induced occupational rhinitis and asthma underestimated?

Amines, some of which are known to cause asthma, are frequently present in the work environment, but are rarely identified as being responsible for occupational rhinitis (OR) or asthma (OA). However, amine-induced OR/OA may be underreported. To discuss this hypothesis, we report a series of patients with positive amine-specific nasal provocation test (NPT). Review of the medical charts of 37 patients with OR (alone or associated with asthma), submitted to a NPT with an aliphatic or alicyclic amine (except for EDTA) present in a product used at work. Most patients worked in the healthcare sector or for a cleaning company. Amines were mostly present in cleaning products. Seven patients had a positive NPT. NPTs were positive for the following amines: bis(aminopropyl)laurylamine, C12-C18 alkyldimethylamine oxides, bis(2-hydroxyethyl)tallowamine oxides, 3-dimethylaminopropylamine, 2,2'-dimethyl-4,4'-methylene-bis(cyclohexylamine), lauryldimethylamine oxide. NPTs were negative for the following amines: monoethanolamine, diethanolamine, triethanolamine, isopropanolamine, triethylamine, triethylenetetramine, aminopropyltriethoxysilane, alkylpropylenediamineguanidine acetate. The frequency of amine-induced OR/OA may be underestimated, particularly when cleaning products are incriminated. Comprehensive investigation of all cases is mandatory to ensure an efficient prevention policy and consequently a good clinical and socio-occupational prognosis of occupational respiratory disease.

Cysteine Residues Impact the Stability and Micelle Interaction Dynamics of the Human Mitochondrial β-Barrel Anion Channel hVDAC-2

The anti-apoptotic 19-stranded transmembrane human voltage dependent anion channel isoform 2 (hVDAC-2) β-barrel stability is crucial for anion transport in mitochondria. The role of the unusually high number of cysteine residues in this isoform is poorly understood. Using a Cys-less construct of hVDAC-2, we haveinvestigated the contribution of cysteines to channel function, barrel stability and its influence on the strength of protein-micelle interactions. We observe that despite the overall preservation in barrel structure upon cysteine mutation, subtle local variations in the mode of interaction of the barrel with its refolded micellar environment arise, which may manifest itself in the channel activity of both the proteins.Fluorescence measurements of the Trp residues in hVDAC-2 point to possible differences in the association of the barrel with lauryldimethylamine oxide (LDAO) micelles. Upon replacement of cysteines in hVDAC-2, our data suggests greater barrel rigidity by way of intra-protein interactions. This, in turn, lowers the equilibrium barrel thermodynamic parameters in LDAOby perturbingthe stability of the protein-micelle complex. In addition to this, we also find a difference in the cooperativity of unfolding upon increasing the LDAO concentration, implying the importance of micelle concentration and micelle-protein ratios on the stability of this barrel. Our results indicate that the nine cysteine residues of hVDAC-2 are the key in establishing strong(er) barrel interactions with its environment and also impart additional malleability to the barrel scaffold.

The Mechanism of Denaturation and the Unfolded State of the α-Helical Membrane-Associated Protein Mistic

In vitro protein-folding studies using chemical denaturants such as urea are indispensible in elucidating the forces and mechanisms determining the stability, structure, and dynamics of water-soluble proteins. By contrast, α-helical membrane-associated proteins largely evade such approaches because they are resilient to extensive unfolding. We have used optical and NMR spectroscopy to provide an atomistic-level dissection of the effects of urea on the structure and dynamics of the α-helical membrane-associated protein Mistic as well as its interactions with detergent and solvent molecules. In the presence of the zwitterionic detergent lauryl dimethylamine oxide, increasing concentrations of urea result in a complex sequence of conformational changes that go beyond simple two-state unfolding. Exploiting this finding, we report the first high-resolution structural models of the urea denaturation process of an α-helical membrane-associated protein and its completely unfolded state, which contains almost no regular secondary structure but nevertheless retains a topology close to that of the folded state.