共查询到20条相似文献,搜索用时 15 毫秒
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P Wittung-Stafshede 《Biochimica et biophysica acta》1999,1432(2):401-405
Both the enthalpic and entropic contributions to unfolding of three heme proteins, cytochrome b(562), cytochrome c and myoglobin, are larger for the reduced than for the oxidized form. Thus, the higher thermodynamic stability of a reduced, as compared to an oxidized, heme protein is the net result of a large increase of favorable enthalpy and a small increase in unfavorable entropy. Upon comparing the unfolding energetics of the heme proteins to those of other single-domain proteins I find that protein length is the primary determinant of the thermodynamics. 相似文献
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The equilibrium unfolding of calf intestinal alkaline phosphatase in guanidinium chloride (GdmCl) solutions was studied by following the fluorescence and ultraviolet difference spectra. At low concentrations of GdmCl (< 1.6 M), the fluorescence intensity decreased with a slight red shift of the emission maximum from 332 nm to 344 nm. An unfolding intermediate state was observed at a broad concentration range of GdmCl as a denaturant (between 1.6 and 2.6 M). This intermediate was characterized by increased fluorescence emission intensity, ultraviolet difference absorption at 236 nm and 260 nm, as well as increased binding to the protein and red shift of the fluorescence probe 1-anilinonaphthalene-8-sulfonic acid. 相似文献
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Tetsuichi Wazawa Takashi Miyazaki Yoshihiro Sambongi Makoto Suzuki 《Biophysical chemistry》2010,151(3):160-2876
Dielectric relaxation (DR) study was performed to reveal the hydration change of Pseudomonas aeruginosa ferric cytochrome c551 (PA c551) in dilute aqueous solutions upon the acid unfolding which undergoes a two-state transition. The DR spectrum of a small spherical region containing a PA c551 molecule and its surrounding water shell was derived from the solution and solvent spectra by dielectric mixture theories. The derived spectrum was well-fitted with a sum of a Debye relaxation component (C1) with a DR frequency around 4.7 GHz and the bulk solvent component (CB). Upon acid unfolding, the DR amplitude of CB decreased with decreasing pH in an inverse manner to that of C1, while the total DR amplitude was almost constant. It indicates that C1 is due to the hydration water of PA c551. Little change in the DR frequency of C1 and a 1.7-fold increase in hydration number were observed. 相似文献
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《FEBS letters》2013,587(8):1081-1088
The transduction of biological signals often involves structural rearrangements of proteins in response to input signals, which leads to functional outputs. This review discusses the role of regulated partial and complete protein unfolding as a mechanism of controlling protein function and the prevalence of this regulatory mechanism in signal transduction pathways. The principles of regulated unfolding, the stimuli that trigger unfolding, and the coupling of unfolding with other well characterized regulatory mechanism are discussed. 相似文献
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The equilibrium unfolding of pig kidney aminoacylase in guanidinium chloride (GdmCl) solutions was studied by following the fluorescence and circular dichroism (CD). At low concentrations of GdmCl, less than 1.0 M, the fluorescence intensity decreased with a slight red shift of the emission maximum (from 335 to 340 nm). An unfolding intermediate was observed in low concentrations of denaturant (between 1.2 and 1.6 M GdmCl). This intermediate was characterized by a decreased fluorescence emission intensity, a red-shifted emission maximum, and increased binding of the fluorescence probe 1-anilino-8-naphthalenesulfonate. No significant changes of the secondary structure were indicated by CD measurement. This conformation state is similar to a molten globule state which may exist in the pathway of protein folding. Further changes in the fluorescence properties occurred at higher concentrations of GdmCl, more than 1.6 M, with a decrease in emission intensity and a significant red shift of the emission maximum from 340 to 354 nm. In this stage, the secondary structure was completely broken. A study of apo-enzyme (Zn2+-free enzyme) produced similar results. However, comparison of the changes of the fluorescence emission spectra of native (Holo-) enzyme with Zn2+-free (Apo-) enzyme at low GdmCl concentrations showed that the structure of the Holo-enzyme was more stable than that of the Apo-enzyme. 相似文献
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The heat capacity change upon unfolding (deltaC(p)) is a thermodynamic parameter that defines the temperature dependence of the thermodynamic stability of proteins; however, physical basis of the heat capacity change is not completely understood. Although empirical surface area-based calculations can predict heat capacity changes reasonably well, accumulating evidence suggests that changes in hydration of those surfaces is not the only parameter contributing to the observed heat capacity changes upon unfolding. Because packing density in the protein interior is similar to that observed in organic crystals, we hypothesized that changes in protein dynamics resulting in increased rigidity of the protein structure might contribute to the observed heat capacity change upon unfolding. Using differential scanning calorimetry we characterized the thermodynamic behavior of a serine protease inhibitor eglin C and two eglin C variants with altered native state dynamics, as determined by NMR. We found no evidence of changes in deltaC(p) in either of the variants, suggesting that changes in rigidity do not contribute to the heat capacity change upon unfolding in this model system. 相似文献
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The folding of an oligomeric protein poses an extra challenge to the folding problem because the protein not only has to fold correctly; it has to avoid nonproductive aggregation. We have carried out over 100 molecular dynamics simulations using an implicit solvation model at different temperatures to study the unfolding of one of the smallest known tetramers, p53 tetramerization domain (p53tet). We found that unfolding started with disruption of the native tetrameric hydrophobic core. The transition state for the tetramer to dimer transition was characterized as a diverse ensemble of different structures using Phi value analysis in quantitative agreement with experimental data. Despite the diversity, the ensemble was still native-like with common features such as partially exposed tetramer hydrophobic core and shifts in the dimer-dimer arrangements. After passing the transition state, the secondary and tertiary structures continued to unfold until the primary dimers broke free. The free dimer had little secondary structure left and the final free monomers were random-coil like. Both the transition states and the unfolding pathways from these trajectories were very diverse, in agreement with the new view of protein folding. The multiple simulations showed that the folding of p53tet is a mixture of the framework and nucleation-condensation mechanisms and the folding is coupled to the complex formation. We have also calculated the entropy and effective energy for the different states along the unfolding pathway and found that the tetramerization is stabilized by hydrophobic interactions. 相似文献
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López-Arenas L Solís-Mendiola S Padilla-Zúñiga J Hernández-Arana A 《Biochimica et biophysica acta》2006,1764(7):1260-1267
We studied the effect of three electrolytes (LiCl, Na(2)SO(4), GuHCl) on the unfolding reaction of chymopapain, a two-domain protein belonging in the papain family of cysteine proteinases. Due to methodological reasons, these studies were carried out at pH 1.5 where the protein unfolds following biphasic kinetics. We have observed the presence of two different effects of electrolyte concentration on the unfolding reactions. At low ionic strength, the ionic atmosphere brought about an increase in reaction rates, regardless of the type of ions being present; this effect is attributed to a general "electrostatic screening" of charge-charge interactions in the macromolecule. At high ionic strength, each electrolyte exerted a distinctively different effect: both rate constants were largely increased by GuHCl (a well-known protein denaturant), but only slightly by LiCl; in contrast, Na(2)SO(4) (a good precipitant) decreased the value of both unfolding rates. These ion-specific (Hofmeister) effects were further used to estimate changes in accessible surface area (DeltaASA) upon formation of the transition states (TS) for unfolding. Results obtained with LiCl and Na(2)SO(4), which we analyzed by means of a parameterization derived from published solubility data of amino acid derivatives, are consistent with DeltaASA increments (for each phase) of about 8.0% of the total theoretical DeltaASA for complete unfolding of the chymopapain molecule. Results in the presence of GuHCl, which were analyzed by using a previous parameterization of protein unfolding data, gave larger DeltaASAs of activation, equivalent to 13 and 16% of the total unfolding DeltaASA. 相似文献
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Reversing the loop lengths of the small protein S6 by circular permutation has a dramatic effect on the transition state structure: it changes from globally diffuse to locally condensed. The phenomenon arises from a biased dispersion of the contact energies. Stability data derived from point mutations throughout the S6 structure show that interactions between residues that are far apart in sequence are stronger than those that are close. This entropy compensation drives all parts of the protein to fold simultaneously and produces the diffuse transition-state structure typical for two-state proteins. In the circular permutant, where strong contacts and short sequence separations are engineered to concur, the transition state becomes atypically condensed and polarized. Taken together with earlier findings that S6 may also fold by a 'collapsed' trajectory with an intermediate, the results suggest that this protein may fold by a multiplicity of mechanisms. The observations indicate that the diffuse transition state of S6 is not required for folding but could be an evolutionary development to optimize cooperativity. 相似文献
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The enthalpies of solvation of 17 amino acids were evaluated by using the sublimation enthalpies of amino acids and the standard enthalpies of their solution in water. An equation was derived, which relates the volume-specific enthalpy of sublimation (deltaH(subl)/V(w)) to the sum of the common bond lengths in molecules (sigman(i)l(i)) of substances examined. The results obtained are interpreted in terms of the effect of hydrophobic and hydrophilic side chain on the interactions between the zwitterions of amino acids and water molecules. 相似文献
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In contrast to single-domain proteins unfolding of larger multi-domain proteins is often irreversible. In a comparative case study on three different multi-domain proteins (phosphoglycerate kinase: PGK and two homologous alpha-amylases: TAKA and BLA) we investigated properties of unfolded states and their ability to fold back into the native state. For this purpose guanidine hydrochloride, alkaline pH, and thermal unfolded states were characterized. Structural alterations upon unfolding and refolding transitions were monitored using fluorescence and CD spectroscopy. Static and dynamic light scattering was employed to follow aggregation processes. Furthermore, proper refolding was also investigated by enzyme activity measurements. While for PGK at least partial reversible unfolding transitions were observed in most cases, we found reversible unfolding for TAKA in the case of alkaline pH and GndHCl induced unfolding. BLA exhibits reversible unfolding only under conditions with high concentrations of protecting osmolytes (glycerol), indicating that aggregation of the unfolded state is the main obstacle to achieve proper refolding for this protein. Structural properties, such as number and size of domains, secondary structure contents and compositions within domains, and domain topology were analyzed and considered in the interpretation of differences in refolding behavior of the investigated proteins. 相似文献
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Keisuke Yagawa Koji Yamano Takaomi Oguro Masahiro Maeda Takehiro Sato Takaki Momose Shin Kawano Toshiya Endo 《Protein science : a publication of the Protein Society》2010,19(4):693-702
Point mutations in proteins can have different effects on protein stability depending on the mechanism of unfolding. In the most interesting case of I27, the Ig‐like module of the muscle protein titin, one point mutation (Y9P) yields opposite effects on protein stability during denaturant‐induced “global unfolding” versus “vectorial unfolding” by mechanical pulling force or cellular unfolding systems. Here, we assessed the reason for the different effects of the Y9P mutation of I27 on the overall molecular stability and N‐terminal unraveling by NMR. We found that the Y9P mutation causes a conformational change that is transmitted through β‐sheet structures to reach the central hydrophobic core in the interior and alters its accessibility to bulk solvent, which leads to destabilization of the hydrophobic core. On the other hand, the Y9P mutation causes a bend in the backbone structure, which leads to the formation of a more stable N‐terminal structure probably through enhanced hydrophobic interactions. 相似文献
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Mechanically unfolding proteins: the effect of unfolding history and the supramolecular scaffold
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Zinober RC Brockwell DJ Beddard GS Blake AW Olmsted PD Radford SE Smith DA 《Protein science : a publication of the Protein Society》2002,11(12):2759-2765
The mechanical resistance of a folded domain in a polyprotein of five mutant I27 domains (C47S, C63S I27)(5)is shown to depend on the unfolding history of the protein. This observation can be understood on the basis of competition between two effects, that of the changing number of domains attempting to unfold, and the progressive increase in the compliance of the polyprotein as domains unfold. We present Monte Carlo simulations that show the effect and experimental data that verify these observations. The results are confirmed using an analytical model based on transition state theory. The model and simulations also predict that the mechanical resistance of a domain depends on the stiffness of the surrounding scaffold that holds the domain in vivo, and on the length of the unfolded domain. Together, these additional factors that influence the mechanical resistance of proteins have important consequences for our understanding of natural proteins that have evolved to withstand force. 相似文献
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Renthal R 《Biochemistry》2006,45(49):14559-14566
Reversible unfolding of helical transmembrane proteins could provide valuable information about the free energy of interaction between transmembrane helices. Thermal unfolding experiments suggest that this process for integral membrane proteins is irreversible. Chemical unfolding has been accomplished with organic acids, but the unfolding or refolding pathways involve irreversible steps. Sodium dodecyl sulfate (SDS) has been used as a perturbant to study reversible unfolding and refolding kinetics. However, the interpretation of these experiments is not straightforward. It is shown that the results could be explained by SDS binding without substantial unfolding. Furthermore, the SDS-perturbed state is unlikely to include all of the entropy terms involved in an unfolding process. Alternative directions for future research are suggested: fluorinated alcohols in homogeneous solvent systems, inverse micelles, and fragment association studies. 相似文献
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J.F. Willart N. Dujardin E. Dudognon F. Danède M. Descamps 《Carbohydrate research》2010,345(11):1613-4426
The possibility to amorphize anhydrous crystalline sugars, like lactose, trehalose and glucose, by mechanical milling was previously reported. We test here the possibility to amorphize the corresponding crystalline hydrates: lactose monohydrate, trehalose dihydrate and glucose monohydrate using fully identical milling procedures. The results show that only the first hydrate amorphizes while the other two remain structurally invariant. These different behaviours are attributed to the plasticizing effect of the structural water molecules which can decrease the glass transition temperature below the milling temperature. The results reveal clearly the fundamental role of the glass transition in the solid-state amorphization process induced by milling, and they also explain why crystalline hydrates are systematically more difficult to amorphize by milling than their anhydrous counterpart. The investigations have been performed by differential scanning calorimetry and powder X-ray diffraction. 相似文献