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The chemical reaction between Fe and lacquer has been used to create the black color in lacquer coatings since ancient times. Here, the effects of Fe ion addition, UV irradiation, and heating on the microscopic structures of black lacquer films were investigated by using X-ray absorption near edge structure (XANES), extended X-ray absorption fine structure (EXAFS), Fourier transform-infrared spectroscopy (FT-IR), small-angle X-ray scattering (SAXS), and small angle neutron scattering (SANS). The EXAFS result indicated that heating and UV irradiation made the coordination structure of Fe3+ in the lacquer nonuniform, and that heating caused the greatest nonuniformity. The FT-IR, SAXS, and SANS results demonstrated that the microscopic structural changes in the black lacquer films were induced by both heating and UV irradiation, but the changes were different. Heating caused a substantial structural change on the nanoscale, and UV irradiation mainly caused changes in the molecular binding mode. The results provide important knowledge for analyzing archeological lacquer samples and for developing lacquer-based materials. This work also demonstrates the utility of the complementary use of XANES, EXAFS, FT-IR, SAXS, and SANS for nondestructive analysis of black lacquer in precious cultural relics.
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The mechanisms of protein stabilization by uncharged solutes, such as polyols and sugars, have been intensively studied with respect to the chemical thermodynamics of molecular crowding. In particular, many experimental and theoretical studies have been conducted to explain the mechanism of the protective action on protein structures by glycerol through the relationship between hydration and glycerol solvation on protein surfaces. We used wide-angle x-ray scattering (WAXS), small-angle neutron scattering, and theoretical scattering function simulation to quantitatively characterize the hydration and/or solvation shell of myoglobin in aqueous solutions of up to 75% v/v glycerol. At glycerol concentrations below â¼40% v/v, the preservation of the hydration shell was dominant, which was reasonably explained by the preferential exclusion of glycerol from the protein surface (preferential hydration). In contrast, at concentrations above 50% v/v, the partial penetration or replacement of glycerol into or with hydration-shell water (neutral solvation by glycerol) was gradually promoted. WAXS results quantitatively demonstrated the neutral solvation, in which the replacement of hydrated water by glycerol was proportional to the volume fraction of glycerol in the solvent multiplied by an exchange rate (ß ≤ 1). These phenomena were confirmed by small-angle neutron scattering measurements. The observed WAXS data covered the entire hierarchical structure of myoglobin, ranging from tertiary to secondary structures. We separately analyzed the effect of glycerol on the thermal stability of myoglobin at each hierarchical structural level. The thermal transition midpoint temperature at each hierarchical structural level was raised depending on the glycerol concentration, with enhanced transition cooperativeness between different hierarchical structural levels. The onset temperature of the helix-to-cross ß-sheet transition (the initial process of amyloid formation) was evidently elevated. However, oligomerization connected to fibril formation was suppressed, even at a low glycerol concentration.
Assuntos
Glicerol/farmacologia , Mioglobina/química , Temperatura , Água/química , Animais , Relação Dose-Resposta a Droga , Conformação Proteica em alfa-Hélice/efeitos dos fármacos , Conformação Proteica em Folha beta/efeitos dos fármacos , Solventes/químicaRESUMO
Alkyl-methyl-imidazolium ionic liquids CnmimX (n: alkyl-carbon number, X: anion) have short-range layer structures consisting of ionic and neutral (alkylchain) domains. To investigate the temperature dependences of the interlayer, interionic group, and inter-alkylchain correlations, we have measured the neutron diffraction (ND) of C16mimPF6, C9.5mimPF6, and C8mimPF6 in the temperature region from 4 K to 470 K. The quasielastic neutron scattering (QENS) of C16mimPF6 was also measured to study the dynamics of each correlation. C16mimPF6 shows a first-order transition between the liquid (L) and liquid crystalline (LC) phases at Tc = 394 K. C8mimPF6 exhibits a glass transition at Tg = 200 K. C9.5mimPF6, which is a 1:3 mixture between C8mimPF6 and C10mimPF6, has both transitions at Tc = 225 K and Tg = 203 K. In the ND experiments, all samples exhibit three peaks corresponding to the correlations mentioned above. The widths of the interlayer peak at ca. 0.2 Å-1 changed drastically at the L-LC transitions, while the interionic peaks at ca. 1 Å-1 exhibited a small jump at Tc. The peak position and area of the three peaks did not change much at the transition. The structural changes were minimal at Tg. The QENS experiments demonstrated that the relaxation time of the interlayer motion increased tenfold at Tc, while those of other motions were monotonous in the whole temperature region. The structural and dynamical changes mentioned above are characteristic of the L-LC transition in imidazolium-based ionic liquids.
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A major form of proteasome activator PA28 is a heteroheptamer composed of interferon-γ-inducible α and ß subunits, which share approximately 50% amino acid identity and possess distinct insert loops. This activator forms a complex with the 20S proteasome and thereby stimulates proteasomal degradation of peptides in an ATP-independent manner, giving rise to smaller antigenic peptides presented by major histocompatibility complex class I molecules. In this study, we performed biophysical and biochemical characterization of the structure and function of the PA28 hetero-oligomer. Deuteration-assisted small-angle neutron scattering demonstrated three α and four ß subunits are alternately arranged in the heptameric ring. In this arrangement, PA28 loops surround the central pore of the heptameric ring (site for peptide entry). Activating the 20S proteasome with a PA28 mutant that lacked the α subunit loops cleaved model substrates longer than a nonapeptide with better efficiency when compared to wild-type PA28. Based on these data, we hypothesize that the flexible PA28 loops act as gatekeepers, which function to select the length of peptide substrates to be transported between the proteolytic chamber and the extra-proteasomal medium.
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Complexo de Endopeptidases do Proteassoma/química , Complexo de Endopeptidases do Proteassoma/metabolismo , Animais , Camundongos , Peptídeos/química , Conformação Proteica , Multimerização Proteica , Subunidades Proteicas/química , Subunidades Proteicas/metabolismo , ProteóliseRESUMO
It is well-known that eutectic gold-silicon (Au-Si) alloys exhibit anomalous melting point depression, which is more than 1000 °C from the melting point of elemental Si (1414 °C). The melting point depression in eutectic alloys is generally explained in terms of a decrease of the free energy by mixing. However, it is difficult to understand the anomalous melting point depression only from the stability of the homogeneous mixing. Some researchers suggest that there are concentration fluctuations in the liquids, where the atoms are inhomogeneously mixed. In this paper, we measure the small-angle neutron scattering (SANS) of Au81.4Si18.6(eutectic composition) and Au75Si25(off-eutectic composition) at temperatures from room temperature to 900 °C in both solid and liquid states to observe such concentration fluctuations directly. It is surprising that large SANS signals are observed in the liquids. This indicates that there are concentration fluctuations in the liquids. The concentration fluctuations are characterized by either the correlation lengths in multiple length scales or surface fractals. This finding yields new insight into the mixing state in the eutectic liquids. The mechanism of the anomalous melting point depression is discussed based on the concentration fluctuations.
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The performance of a polymer electrolyte fuel cell can be enhanced by improving the proton conductivity of the catalyst layer, where the oxygen reduction reaction generates electrochemical power. Protons are conducted through the ionomer coatings on catalyst-supporting carbon particles, which form porous structures that facilitate oxygen diffusion during the reaction within the catalyst layer. Therefore, while a higher ionomer content in the catalyst layer is favorable, the proton conductivity is additionally governed by the type of carbon support. As the influence of the ionomer distribution is not fully understood, we introduce a novel proton conductivity model for use in simulating catalyst layers with various amounts of ionomers and different carbon types. This proton conductivity model considers that several ionomers occur as thin films with drastically suppressed proton conductivities. Although evaluating the thin-film ionomer fraction is challenging, proton-conducting ion clusters in thick-film ionomers have been detected by characterizing the catalyst layers via small-angle neutron scattering. Our model reveals that reducing the fraction of the thin-film ionomer or avoiding factors that suppress its proton conduction improves the performance of the catalyst layer.
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Semi-crystalline polymers exhibit microphase separation into crystalline and amorphous domains characterized by multiple structural levels with sizes ranging from ångströms to hundreds of nanometres. The combination of small-angle (SANS) and wide-angle (WANS) neutron scattering on the same beamline enables reliable in situ characterization of such materials under application-relevant conditions, with the unique advantage of contrast variation by controlled labelling, allowing the structure of such multi-component systems to be resolved in detail. This paper reports a structural analysis performed on deuterated polymer membranes based on syndiotactic polystyrene (sPS) using an extended Q-range SANS and WANS combination, always with the same neutron scattering instrument, either a pinhole SANS diffractometer installed at a research reactor or a 'small- and wide-angle' time-of-flight diffractometer installed at a neutron spallation source. sPS is a semi-crystalline material that becomes hydrophilic and proton conducting when suitable functionalization is achieved by thin film sulfonation, and can form various co-crystalline complexes (clathrates) with small organic molecules stored in the crystalline phase as guests in the vacancies between the polymer helices. Therefore, this material is interesting not only for its conducting properties but also for its versatility as a model system to evaluate the usefulness of extended Q-range neutron scattering in such studies. Variation of neutron contrast was achieved in the amorphous hydrophilic phase by using H2O or D2O to hydrate the membranes and in the crystalline phase by loading the clathrates with deuterated or protonated guest molecules. The experimental approach, the advantages and limitations of the two types of instrumentation used in such analyses, and the main results obtained with respect to the structural characterization of sulfonated sPS membranes under different hydration and temperature conditions are reported, and the potential of this method for similar structural studies on other semi-crystalline polymeric materials is discussed.
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When functionalized by the solid-state sulfonation process, the amorphous regions of the semi-crystalline syndiotactic-polystyrene (sPS) become hydrophilic, and thus can conduct protons upon membrane hydration, which increases the interest in this material as a potential candidate for applications with proton exchange membranes. The resistance of sulfonated sPS to oxidative decomposition can be improved by doping the membrane with fullerenes. In previous work, we have described the morphology in hydrated sulfonated sPS films doped with fullerenes on different length scales as determined by small-angle neutron scattering (SANS) and the structural changes in such membranes as a function of the degree of hydration and temperature. In the current work, we report on the relationship between the morphology of hydrated domains as obtained by SANS and the proton conductivity in sulfonated sPS-fullerene composite membranes at different temperature and relative humidity (RH) conditions. Based on this combined experimental approach, clear evidence for the formation and evolution of the hydrated domains in functionalized sPS membranes has been provided and a better understanding of the hydration and conductivity pathways in this material has been obtained.
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Dynamically crosslinked gels are appealing materials for applications that require time-dependent mechanical responses. DNA duplexes are ideal crosslinkers for building such gels because of their excellent sequence addressability and flexible tunability in bond energy. However, the mechanical responses of most DNA gels are complicated and unpredictable. Here, a DNA gel with a highly homogeneous gel network and well predictable mechanical behaviors is demonstrated by using a pair of star-polymer-DNA precursors with presimulated DNA sequences showing the two-state transition. The melting curve analysis of the DNA gels reveals the good correspondence between the thermodynamic potentials of the DNA crosslinkers and the presimulated values by DNA calculators. Stress-relaxation tests and dissociation kinetics measurements show that the macroscopic relaxation time of the DNA gels is approximately equal to the lifetime of the DNA crosslinkers over 4 orders of magnitude from 0.1-2000 s. Furthermore, a series of durability tests find the DNA gels are hysteresis-less and self-healable after the applications of repeated temperature and mechanical stimuli. These results demonstrate the great potential of star-polymer-DNA precursors for building gels with predictable and tunable viscoelastic properties, suitable for applications such as stress-response extracellular matrices, injectable solids, and soft robotics.
Assuntos
DNA , Polímeros , Géis/química , Polímeros/química , Temperatura , TermodinâmicaRESUMO
The ionomers distributed on carbon particles in the catalyst layer of polymer electrolyte fuel cells (PEFCs) govern electrical power via proton transport and oxygen permeation to active platinum. Thus, ionomer distribution is a key to PEFC performance. This distribution is characterized by ionomer adsorption and deposition onto carbon during the catalyst-ink coating process; however, the adsorbed and deposited ionomers cannot easily be distinguished in the catalyst layer. Therefore, we identified these two types of ionomers based on the positional correlation between the ionomer and carbon particles. The cross-correlation function for the catalyst layer was obtained by small-angle neutron scattering measurements with varying contrast. From fitting with a model for a fractal aggregate of polydisperse core-shell spheres, we determined the adsorbed-ionomer thickness on the carbon particle to be 51 Å and the deposited-ionomer amount for the total ionomer to be 50%. Our technique for ionomer differentiation can be used to optimally design PEFC catalyst layers.
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The present study aimed to investigate the thermal- and pH-dependent gelation behavior of gelatin/HPMCP blends using ultraviolet (UV) spectrophotometry, viscoelasticity, and dynamic light scattering (DLS). We found that the release of lisinopril from gelatin/HPMCP gels can be inhibited at low pH. UV spectrophotometric analysis showed that pH had a significant effect on the transparency of aqueous HPMCP systems and gelatin/HPMCP gels. The viscoelastic patterns of gelatin/HPMCP at pH 4.6 considerably differed from those of gelatin/HPMCP at pH 5.2 and 6.0. DLS measurements showed that HPMCP molecules in low concentrations underwent strong aggregation below pH 4.8. Such HPMCP aggregation induces a physical barrier in the matrix structures of the gelatin/HPMCP gels, which inhibits the drug release at pH 1.2. This hydrogel delivery system using polymer blends of gelatin/HPMCP can be used in oral gel formulations with pH-responsive properties.
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Liberação Controlada de Fármacos , Gelatina/química , Metilcelulose/análogos & derivados , Difusão Dinâmica da Luz , Módulo de Elasticidade , Concentração de Íons de Hidrogênio , Lisinopril/farmacologia , Metilcelulose/química , Polímeros/química , Reologia , Eletricidade EstáticaRESUMO
Amyloids are implicated in many diseases, and disruption of lipid membrane structures is considered as one possible mechanism of pathology. In this paper we investigate interactions between an aggregating peptide and phospholipid membranes, focusing on the nanometer-scale structures of the aggregates formed, as well as on the effect on the aggregation process. As a model system, we use the small amyloid-forming peptide named NACore, which is a fragment of the central region of the protein α-synuclein that is associated with Parkinson's disease. We find that phospholipid vesicles readily associate with the amyloid fibril network in the form of highly distorted and trapped vesicles that also may wet the surface of the fibrils. This effect is most pronounced for model lipid systems containing only zwitterionic lipids. Fibrillation is found to be retarded by the presence of the vesicles. At the resolution of our measurements, which are based mainly on cryogenic transmission electron microscopy (cryo-TEM), X-ray scattering, and circular dichroism (CD) spectroscopy, we find that the resulting aggregates can be well fitted as linear combinations of peptide fibrils and phospholipid bilayers. There are no detectable effects on the cross-ß packing of the peptide molecules in the fibrils, or on the thickness of the phospholipid bilayers. This suggests that while the peptide fibrils and lipid bilayers readily co-assemble on large length-scales, most of them still retain their separate structural identities on molecular length-scales. Comparison between this relatively simple model system and other amyloid systems might help distinguish aspects of amyloid-lipid interactions that are generic from aspects that are more protein specific. Finally, we briefly consider possible implications of the obtained results for in-vivo amyloid toxicity.
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The interaction of nanoparticles with surfactants is extensively used in a wide range of applications from enhancing colloidal stability to phase separation processes as well as in the synthesis of noble functional materials. The interaction is highly specific depending on the charged nature of the surfactant. In the case of nonionic surfactants, the micelles adsorb on the surface of nanoparticles. The adsorption of nonionic surfactant C12E10 as a function of surfactant concentration for two different sizes of anionic silica nanoparticles (16 and 27 nm) has been examined using dynamic light scattering (DLS) and small-angle neutron scattering (SANS). SANS measurements have been carried out under different contrast-matched conditions, where nanoparticles, as well as surfactant micelles, have been contrast-matched to the solvent. The adsorption of micelles is determined from the contrast-matched condition of silica nanoparticles with the solvent. SANS data under surfactant contrast-matched condition suggest that there is no modification in the structure and/or interaction of the silica nanoparticles in presence of nonionic micelles. The adsorption of micelles on nanoparticles is found to follow an exponential behavior with respect to the surfactant concentration. These results are consistent with the variation of hydrodynamic size of nanoparticle-surfactant system in DLS. The study on different-sized nanoparticles shows that the lower curvature enhances the packing fraction whereas the loss of surface-to-volume ratio suppresses the fraction of adsorbed micelles with the increase in the nanoparticle size. The adsorption coefficient has higher value for the larger size of the nanoparticles. In the mixed system of two sizes of nanoparticles, no preferential selectivity of micelle adsorption is observed.
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Membranes based on sulfonated synditoactic polystyrene (s-sPS) were thoroughly characterized by contrast variation small-angle neutron scattering (SANS) over a wide Q-range in dry and hydrated states. Following special sulfonation and treatment procedures, s-sPS is an attractive material for fuel cells and energy storage applications. The film samples were prepared by solid-state sulfonation, resulting in uniform sulfonation of only the amorphous phase while preserving the crystallinity of the membrane. Fullerenes, which improve the resistance to oxidation decomposition, were incorporated in the membranes. The fullerenes seem to be chiefly located in the amorphous regions of the samples, and do not influence the formation and evolution of the morphologies in the polymer films, as no significant differences were observed in the SANS patterns compared to the fullerenes-free s-sPS membranes, which were investigated in a previous study. The use of uniaxially deformed film samples, and neutron contrast variation allowed for the identification and characterization of different structural levels with sizes between nm and µm, which form and evolve in both the dry and hydrated states. The scattering length density of the crystalline regions was varied using the guest exchange procedure between different toluene isotopologues incorporated into the sPS lattice, while the variation of the scattering properties of the hydrated amorphous regions was achieved using different H2O/D2O mixtures. Due to the deformation of the films, the scattering characteristics of different structures can be distinguished on specific detection sectors and at different detection distances after the sample, depending on their size and orientation.
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HYPOTHESIS: Gemini surfactant 12-2-12 (dimethylene-1,2-bis(dodecyl dimethylammonium bromide)) solutions are known to show shear thickening and thinning under salt-free conditions. Because the rheological behavior of the wormlike micelles formed by 12-2-12 in solution is related to their structure, we expected that changes to the precursor structure would affect their rheological behaviors. It is also important to understand the effect of the introduction of a spacer group in the 12-2-12 molecules on the rheological behavior and structure of the wormlike micelles under shear flow. EXPERIMENTS: Simultaneous small-angle neutron scattering and rheological measurements (Rheo-SANS) of the 12-2-12 solutions were performed. We exhaustively studied the structural changes in the wormlike micelles upon increasing shear rate. FINDINGS: We found that the wormlike micelles were oriented in the direction of the flow due to elongation and that changes to the precursor of the wormlike micelles did not affect the shear thickening. As a precursor structural change of shear thinning, the wormlike micelles elongated while maintaining their orientation. We found that an increase in the molecular curvature of the 12-2-12 due to the introduction of a spacer-group contributed to the unusual rheological behaviors of the wormlike micelles in a solution under shear flow.
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The interior of living cells is a molecular-crowding environment, where large quantities of various molecules coexist. Investigations into the nature of this environment are essential for an understanding of both the elaborate biological reactions and the maintenance of homeostasis occurring therein. The equilibrium states of biological macromolecular systems are affected by molecular-crowding environments unmatched by in vitro diluted environments; knowledge about crowding effects is still insufficient due to lack of relevant experimental studies. Recent developments in the techniques of in-cell NMR and large-scale molecular dynamics simulation have provided new insights into the structure and dynamics of biological molecules inside the cells. This study focused on a new experimental technique to directly observe the structure of a specific protein or membrane in condensed crowder solutions using neutron scattering. Deuterated whole-cell debris was used to reproduce an environment that more closely mimics the interior of living cells than models used previously. By the reduction of the background scattering from large amounts of cell debris, we successfully extracted structure information for both small globular protein and small unilamellar vesicle (SUV) from the concentrated cell-debris solution up to a weight ratio of 1:60 for protein/crowder and 1:40 for SUV/crowder.
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Membrana Celular/química , Membrana Celular/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Simulação de Dinâmica Molecular , Difração de NêutronsRESUMO
Small angle neutron scattering (SANS) is a versatile and convenient method to investigate the higher order structure of molecular assembly systems. However, the more complicated a system of interest, the more difficult the interpretation in the SANS profile. In order to increase the reliability of structural analysis on a complicated system, it is desirable to obtain different kinds of structural information from the same sample simultaneously. Polarized infrared spectroscopy is able to provide information about the molecular structure, concentration, and orientation of each chemical species in a system. In order to utilize these advantages of polarized infrared spectroscopy, a simultaneous measurement system was built by incorporating a Fourier transform infrared (FTIR) spectrometer into a time-of-flight small angle neutron scattering instrument covering a wide Q range. Using this system, simultaneous measurements of wide- and small-angle neutron scattering and polarized FTIR spectroscopy was realized for the first time.
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The protective action of sugars against the denaturation of myoglobin was clarified by X-ray and neutron scattering methods. Different types of sugars such as disaccharides (trehalose, sucrose) and monosaccharides (glucose, fructose) were used. Experimental data and theoretical simulation based on three different solvation models (preferential solvation model, nonpreferential solvation model, and preferential exclusion (hydration) model) indicated that sugar molecules were preferentially or weakly excluded from the protein surface and preserved the native protein hydration shell. This trend was more evident for disaccharides. The preferential exclusion shifted gradually to the nonpreferential solvation at higher sugar concentrations. On the protective actions of the sugars against the guanidinium-chloride-mediated denaturation, all sugars, starting from the low concentration of 5% w/v, showed the protective trend toward the protein native structure, especially for the secondary structure. The thermal structural transition temperature of myoglobin was raised by about 4-5 °C, accompanied by amyloid formation, for all hierarchical structural levels. In particular, the oligomer formation of the amyloid aggregates was more suppressed. The above protective action was sugar-dependent. The present results clearly suggest that sugars intrinsically protect the native structure of proteins against chemical and thermal denaturation through the preservative action of the hydration shell.
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Mioglobina/química , Açúcares/química , Animais , Densitometria , Guanidina/química , Cavalos , Difração de Nêutrons , Desnaturação Proteica , Multimerização Proteica , Espalhamento de Radiação , Temperatura , Difração de Raios XRESUMO
Organisms having tolerances against extreme environments produce and accumulate stress proteins and/or sugars in cells against the extreme environment such as high or low temperature, drying, and so forth. Sugars and/or polyols are known to prevent protein denaturation and enzyme deactivation. In particular, trehalose has received considerable attention because of its association with cryptobiosis and anhydrobiosis. This study focuses on the restoration of acid-denatured amyloid transition of myoglobin by trehalose. Myoglobin is known to proceed amyloidogenic reaction under denaturation conditions. We found that acid-denatured myoglobin at an initial process of amyloidogenic reaction (helix-to-sheet transition followed by oligomerization) at 25 °C was substantially restored to its native structure by trehalose. This action was prominent during the early stage of amyloid formation. Recent results showed that sugars are preferentially excluded from the protein surface to preserve its hydration shell and stabilize the protein structure against chemical and thermal denaturation. Therefore, the present results suggest that trehalose will restore the tightly bound water molecules around the hotspot (G-helix) of myoglobin on the amyloid transition by its intrinsic preservative action of the native hydration shell against denaturation. The present finding on the restorative action by trehalose could provide new insights into protein folding and amyloidosis.