Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Protein Denaturation”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 109 records · Page 6Linked to original sources
PubMed2002

Effect of antioxidants, citrate, and cryoprotectants on protein denaturation and texture of frozen cod (Gadus morhua).

To investigate the role of antioxidants and cryoprotectants in minimizing protein denaturation in frozen lean fish, cod fillets were treated with either antioxidants (vitamin C (500 mg kg(-1)) or vitamin C (250 mg kg(-1)) + vitamin E (250 mg kg(-1))), antioxidants (vitamins C + E 250 mg kg(-1)each) with citrate (100 mg kg(-1)), cryoprotectants (4% (w/w) sucrose + 4% (w/w) sorbitol), or a mixture of antioxidants (vitamins C + E 250 mg kg(1)), citrate (100 mg kg(-1)), and cryoprotectants (sucrose 40 g kg(-1) + sorbitol 40 g kg(-1)). Untreated and treated fish samples were stored at -10 degrees C; cod fillets stored at -30 degrees C were used as a control. Stored frozen samples were analyzed at intervals for up to 210 days for changes in protein extractability, thermodynamic parameters (transition temperature T(m) and enthalpy DeltaH), structure by FT-Raman spectroscopy, and rheological properties by large and small deformation tests. Results indicated that protein denaturation and texture changes were minimized in the presence of cryoprotectants, as well as in the presence of antioxidants with citrate, antioxidants alone, or the mixture of antioxidants, citrate, and cryoprotectants. In the presence of increased formaldehyde levels in fish treated with vitamin C, toughening was still lower compared to that of the -10 degrees C control due to the antioxidant property of vitamin C. Thus, ice crystal formation and lipid oxidation products are the major factors that cause protein denaturation in lean frozen fish, and antioxidants in addition to cryoprotectants can be used to minimize toughness.

Animals↗
PubMed1996

Kinetics of adsorption of denatured protein at alumina-water interface.

The kinetics of adsorption of soluble denatured protein, gelatin has been studied at the alumina-water interface as a function of protein concentration, pH, temperature and ionic strength. The rate of adsorption of gelatin has been compared with rate of adsorption of BSA denatured by 8 M urea or 0.05 M SDS. The initial stage for the adsorption process is diffusion-controlled and the surface diffusion coefficients evaluated from equations of Ward and Tordai and by Bull for globular and denatured proteins are found to be widely different from each other. The kinetic data for gelatin fit into a first order rate equation with two rate constants, k1a and k2a. Using Arrhenius equation, the activation energies delta E1* and delta E2* have been evaluated from the values of k1a and k2a respectively. The corresponding changes in values of enthalpy of activation (delta H*), entropy of activation (delta S*) and free energy of activation (delta G*) have been evaluated using Eyring's equation for absolute reaction rate. It has been found that for both gelatin and denatured BSA, in the first kinetic step delta H1* > T delta S1* and for the second step T delta S2* > delta H2.

Adsorption↗
PubMed1996

Reactivation of denatured proteins by 23S ribosomal RNA: role of domain V.

Escherichia coli ribosome, its 50S subunit, or simply the 23S rRNA can reactivate denatured proteins in vitro. Here we show that protein synthesis inhibitors chloramphenicol and erythromycin, which bind to domain V of 23S rRNA of E. coli, can inhibit reactivation of denatured pig muscle lactate dehydrogenase and fungal glucose-6-phosphate dehydrogenase by 23S rRNA completely. Oligodeoxynucleotides complementary to two regions within domain V (which cover sites of chloramphenicol resistant mutations and the putative A site of the incoming aminoacyl tRNA), but not to a region outside of domain V, also can inhibit the activity. Domain V of 23S rRNA, therefore, appears to play a crucial role in reactivation of denatured proteins.

Animals↗
PubMed2000

Trehalose is required for conformational repair of heat-denatured proteins in the yeast endoplasmic reticulum but not for maintenance of membrane traffic functions after severe heat stress.

Saccharomyces cerevisiae cells grown at physiological temperature 24 degrees C require preconditioning at 37 degrees C to acquire tolerance towards brief exposure to 48-50 degrees C. During preconditioning, the cytosolic trehalose content increases remarkably and in the absence of trehalose synthesis yeast cannot acquire thermotolerance. It has been speculated that trehalose protects proteins and membranes under environmental stress conditions, but recently it was shown to assist the Hsp104 chaperone in refolding of heat-damaged proteins in the yeast cytosol. We have demonstrated that heat-denatured proteins residing in the endoplasmic reticulum (ER) also can be refolded once the cells are returned to physiological temperature. Unexpectedly, not only ER chaperones but also the cytosolic Hsp104 chaperone is required for conformational repair events in the ER lumen. Here we show that trehalose facilitates refolding of glycoproteins in the ER after severe heat stress. In the absence of Tps1p, a subunit of trehalose synthase, refolding of heat-damaged glycoproteins to bioactive and secretion-competent forms failed or was retarded. In contrast, membrane traffic operated many hours after severe heat stress even in the absence of the TPS1 gene, demonstrating that trehalose had no role in thermoprotection of membranes engaged in vesicular traffic. However, cytosolic proteins were aggregated and protein synthesis abolished, resulting finally in cell death.

Cell Membrane↗
PubMed1995

[The functional state of denatured proteins: the principles of modelling and the first results].

The aqueous environment near cell membranes should be characterized by a local low pH value (due to attraction of protons by the membrane electrostatic potential) and by a local dielectric constant value (due to proximity of the water-organic boundary). It was proposed to simulate the behaviour of proteins near membranes by their behaviour in water-alcohol mixtures at moderately low pH values. Cytochrome c and retinol-binding protein are shown to be denatured at the values of pH and dielectric constant typical for the aqueous environment near the membrane surfaces. The conformational state of both these proteins under mentioned conditions is similar to the molten globule state. The basic function of the retinol-binding protein (transfer and release of vitamin A) is tightly associated with the transition of this carrier protein into the molten globule-like state, as in this case the release of retinol and protein denaturation are coupled and represent a common cooperative process.

Animals↗
PubMed1998

Modeling of irreversible thermal protein denaturation at varying temperature. I. The model involving two consecutive irreversible steps.

The model of thermal protein denaturation involving two consecutive irreversible steps has been analyzed. The equation describing the dependence of excess heat capacity (Cpex) on temperature (T) under a constant scanning rate has been obtained and used for construction of theoretical Cpex versus T profiles. The change in the shape of theoretical profiles at varied Arrhenius equation parameters, ratio for enthalpies of the first and second steps, and the scanning rate have been studied. It has been shown that formal use of one-step model does not provide adequate information about kinetic features of the process in the case of protein denaturation following the mechanism involving two consecutive irreversible steps.

Animals↗
PubMed1992

Origins and consequences of ligand-induced multiphasic thermal protein denaturation.

The presence of subsaturating levels of a high-affinity ligand has been demonstrated both by experiment and calculation to have far-reaching consequences on thermally induced protein denaturation due to the coupling between the protein denaturation and ligand-binding equilibria. Under such circumstances, a protein may undergo biphasic denaturation even though in the absence of ligand it exhibits a thermogram comprised of a single essentially symmetric endotherm. Up to now, the presence of just 2 maxima in the thermogram has been presented merely as an experimental observation or as the result of equilibrium computations. Here we develop a thermodynamic description of the linkage between these equilibria in which the number of cusps present in the thermogram correlates with the number of resolved steps in the plot of saturation level of remaining native protein vs temperature (i.e., the thermal binding curve). During thermally induced denaturation, the concentration of native protein decreases; thus, the native protein in effect is titrated with ligand at constant total ligand concentration. The free ligand concentration The free ligand concentration increases substantially through the release of bound ligand by unfolding protein thereby increasing the saturation level of the remaining native protein. The form of this thermal binding curve is a function of the number of ligand-binding sites on the protein, the magnitudes of the association constants, and the total ligand and total protein concentrations. As a result, the model predicts multiphasic denaturation of a single cooperative unit when the thermal binding curve consists of discrete multiple steps. The presence of only 2 maxima (i.e., a single cusp) in a thermogram for a protein with multiple sites on the native species derives from the form of the thermal binding curve, which in this case is a single-step sigmoidal plot, and not from the predominant denaturation of unliganded and fully liganded native species. In addition, it is shown that, in general, the contributions from the denaturation of individual native protein species are decidedly non-two-state in character; thus, simple deconvolution should not be carried out. The effects of nonzero values of delta Cp and d delta Cp/dT for denaturation and of changes in enthalpy and in heat capacity for ligand binding, as well as the interaction of ligand with the denatured protein, are explored also.

Binding Sites↗
PubMed2005

Denatured proteins and early folding intermediates simulated in a reduced conformational space.

Conformations of globular proteins in the denatured state were studied using a high-resolution lattice model of proteins and Monte Carlo dynamics. The model assumes a united-atom and high-coordination lattice representation of the polypeptide conformational space. The force field of the model mimics the short-range protein-like conformational stiffness, hydrophobic interactions of the side chains and the main-chain hydrogen bonds. Two types of approximations for the short-range interactions were compared: simple statistical potentials and knowledge-based protein-specific potentials derived from the sequence-structure compatibility of short fragments of protein chains. Model proteins in the denatured state are relatively compact, although the majority of the sampled conformations are globally different from the native fold. At the same time short protein fragments are mostly native-like. Thus, the denatured state of the model proteins has several features of the molten globule state observed experimentally. Statistical potentials induce native-like conformational propensities in the denatured state, especially for the fragments located in the core of folded proteins. Knowledge-based protein-specific potentials increase only slightly the level of similarity to the native conformations, in spite of their qualitatively higher specificity in the native structures. For a few cases, where fairly accurate experimental data exist, the simulation results are in semiquantitative agreement with the physical picture revealed by the experiments. This shows that the model studied in this work could be used efficiently in computational studies of protein dynamics in the denatured state, and consequently for studies of protein folding pathways, i.e. not only for the modeling of folded structures, as it was shown in previous studies. The results of the present studies also provide a new insight into the explanation of the Levinthal's paradox.

Animals↗
PubMed1993

A general method for assigning NMR spectra of denatured proteins using 3D HC(CO)NH-TOCSY triple resonance experiments.

A general approach for assigning the resonances of uniformly 15N- and 13C-labeled proteins in their unfolded state is presented. The assignment approach takes advantage of the spectral dispersion of the amide nitrogen chemical shifts in denatured proteins by correlating side chain and backbone carbon and proton frequencies with the amide resonances of the same and adjacent residues. The 1H resonances of the individual amino acid spin systems are correlated with their intraresidue amide in a 3D 15N-edited 1H,1H-TOCSY-HSQC experiment, which allows the spin systems to be assigned to amino acid type. The spin systems are then linked to the adjacent i-1 spin system using the 3D H(C)(CO)NH-TOCSY experiment. Complete 13C assignments are obtained from the 3D (H)C(CO)NH-TOCSY experiment. Unlike other methods for assigning denatured proteins, this approach does not require previous knowledge of the native state assignments or specific interconversion rates between the native and denatured forms. The strategy is demonstrated by assigning the 1H, 13C, and 15N resonances of the FK506 binding protein denatured in 6.3 M urea.

Algorithms↗
PubMed1983

Studies on the physical state of water in living cells and model systems. V. The warming exothermic reaction of frozen aqueous solution of polyvinylpyrrolidone, poly(ethylene oxide), and urea-denatured proteins.

The warming exothermic peaks (WEX) first described by Luyet and coworkers in solutions of ethylene glycol, glycerol, and polyvinylpyrrolidone(PVP) was interpreted on the basis of a devitrification phenomenon, in which rapid cooling converts normal water to amorphous ice, which devitrifies on warming to the state of normal Ice I. Our present studies of the WEX phenomenon in aqueous solutions of urea-denatured proteins, PVP, PVME, and two types of poly(ethylene oxide) led us to a different conclusion: WEX is the result of a transformation of chilled water in the state of polarized multilayers to Ice I. Among the evidence for this is the fact that WEX of the polymer-water system is indifferent to the rate of cooling between 320 degrees/min. and 1.25 degrees/min. and that polymer-water systems cooled to -70 degrees C remain viscous liquids and not (solid) vitreous ice. The transformation from water dominated by PVP, PVME, and urea-denatured proteins to Ice I can occur rapidly during warming or more slowly when the chilled polymer-water systems are kept at 223 degrees K. To effect this transformation the polymers themselves must undergo movements which tend to be retarded most by the presence of bulky side chains, as in urea-denatured proteins, and least in polymers with no side chains (e.g., Polyox WSR-N750).

Animals↗