Folded State (folded + state)

Distribution by Scientific Domains


Selected Abstracts


The conformational stability of the Streptomyces coelicolor histidine-phosphocarrier protein

FEBS JOURNAL, Issue 11 2004
Characterization of cold denaturation, protein interactions
Thermodynamic parameters describing the conformational stability of the histidine-containing phosphocarrier protein from Streptomyces coelicolor, scHPr, have been determined by steady-state fluorescence measurements of isothermal urea-denaturations, differential scanning calorimetry at different guanidinium hydrochloride concentrations and, independently, by far-UV circular dichroism measurements of isothermal urea-denaturations, and thermal denaturations at fixed urea concentrations. The equilibrium unfolding transitions are described adequately by the two-state model and they validate the linear free-energy extrapolation model, over the large temperature range explored, and the urea concentrations used. At moderate urea concentrations (from 2 to 3 m), scHPr undergoes both high- and low-temperature unfolding. The free-energy stability curves have been obtained for the whole temperature range and values of the thermodynamic parameters governing the heat- and cold-denaturation processes have been obtained. Cold-denaturation of the protein is the result of the combination of an unusually high heat capacity change (1.4 ± 0.3 kcal·mol,1·K,1, at 0 m urea, being the average of the fluorescence, circular dichroism and differential scanning calorimetry measurements) and a fairly low enthalpy change upon unfolding at the midpoint temperature of heat-denaturation (59 ± 4 kcal·mol,1, the average of the fluorescence, circular dichroism and differential scanning calorimetry measurements). The changes in enthalpy (m,Hi), entropy (m,Si) and heat capacity (m,Cpi), which occur upon preferential urea binding to the unfolded state vs. the folded state of the protein, have also been determined. The m,Hi and the m,Si are negative at low temperatures, but as the temperature is increased, m,Hi makes a less favourable contribution than m,Si to the change in free energy upon urea binding. The m,Cpi is larger than those observed for other proteins; however, its contribution to the global heat capacity change upon unfolding is small. [source]


The refolding of type II shikimate kinase from Erwinia chrysanthemi after denaturation in urea

FEBS JOURNAL, Issue 8 2002
Eleonora Cerasoli
Shikimate kinase was chosen as a convenient representative example of the subclass of ,/, proteins with which to examine the mechanism of protein folding. In this paper we report on the refolding of the enzyme after denaturation in urea. As shown by the changes in secondary and tertiary structure monitored by far UV circular dichroism (CD) and fluorescence, respectively, the enzyme was fully unfolded in 4 m urea. From an analysis of the unfolding curve in terms of the two-state model, the stability of the folded state could be estimated as 17 kJ·mol,1. Approximately 95% of the enzyme activity could be recovered on dilution of the urea from 4 to 0.36 m. The results of spectroscopic studies indicated that refolding occurred in at least four kinetic phases, the slowest of which (k = 0.009 s,1) corresponded with the regain of shikimate binding and of enzyme activity. The two most rapid phases were associated with a substantial increase in the binding of 8-anilino-1-naphthalenesulfonic acid with only modest changes in the far UV CD, indicating that a collapsed intermediate with only partial native secondary structure was formed rapidly. The relevance of the results to the folding of other ,/, domain proteins is discussed. [source]


Intrastrand foldamer crosslinking by reductive amination

JOURNAL OF POLYMER SCIENCE (IN TWO SECTIONS), Issue 4 2010
Ronald A. Smaldone
Abstract A series of m -phenylene ethynylene (mPE) foldamers were crosslinked in their helical conformation using a reductive amination-based strategy. This was accomplished by placing aldehyde moieties in the backbone of the oligomer at specific residues, which allowed a diamine crosslinker to covalently link the helical loops together. Three different foldamers with crosslinks placed at different locations in the backbone were synthesized and characterized by mass spectrometry, 1H NMR, and gel permeation chromatography. The effect of the crosslinking on the stability of the folded state was evaluated through solvent denaturation studies. These studies show a reduction in the oligomer's ability to unfold of up to 30% relative to an unmodified mPE oligomer of the same length in solvents that promote unfolding. © 2010 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 48: 927,935, 2010 [source]


Self-assembly of supramolecular polymers into tunable helical structures

JOURNAL OF POLYMER SCIENCE (IN TWO SECTIONS), Issue 6 2008
Ho-Joong Kim
Abstract There is growing interest in the design of synthetic molecules that are able to self-assemble into a polymeric chain with compact helical conformations, which is analogous to the folded state of natural proteins. Herein, we highlight supramolecular approach to the formation of helical architectures and their conformational changes driven by external stimuli. Helical organization in synthetic self-assembling systems can be achieved by the various types of noncovalent interactions, which include hydrogen bonding, solvophobic effects, and metal-ligand interactions. Since the external environment can have a large influence on the strength and configuration of noncovalent interactions between the individual components, stimulus-induced alterations in the intramolecular noncovalent interactions can result in dynamic conformational change of the supramolecular helical structure thus, driving significant changes in the properties of the materials. Therefore, these supramolecular helices hold great promise as stimuli-responsive materials. © 2008 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 46: 1925,1935, 2008 [source]


chicken villin headpiece subdomain;

MAGNETIC RESONANCE IN CHEMISTRY, Issue 9 2009
cross-correlated relaxation;
We have investigated slow correlated motions of neighboring carbonyl and nitrogen nuclei in the backbone of chicken villin headpiece subdomain. Cross-correlated chemical shift modulation experiments were performed at three temperatures where the protein remains in its folded state. The results at 8 °C demonstrate the presence of microseconds to milliseconds timescale motions for a number of residues belonging both to helices and unstructured regions. As the temperature is raised, the motions become progressively less visible. The reduction of the contributions of slow motions into the cross-correlated relaxation rate with the rise in temperature is caused by the increase of the chemical exchange rate constants for the slow motion processes. Copyright © 2009 John Wiley & Sons, Ltd. [source]


The acid-induced folded state of Sac7d is the native state

PROTEIN SCIENCE, Issue 10 2000
Jennifer L. Bedell
Abstract Sac7d unfolds at low pH in the absence of salt, with the greatest extent of unfolding obtained at pH 2. We have previously shown that the acid unfolded protein is induced to refold by decreasing the pH to 0 or by addition of salt (McCrary BS, Bedell J, Edmondson SP, Shriver JW, 1998, J Mol Biol 276:203,224). Both near-ultraviolet circular dichroism spectra and ANS fluorescence enhancements indicate that the acid- and salt-induced folded states have a native fold and are not molten globular. 1H, 15N heteronuclear single quantum coherence NMR spectra confirm that the native, acid-, and salt-induced folded states are essentially identical. The most significant differences in amide 1H and 15N chemical shifts are attributed to hydrogen bonding to titrating carboxyl side chains and through-bond inductive effects. The 1H NMR chemical shifts of protons affected by ring currents in the hydrophobic core of the acid- and salt-induced folded states are identical to those observed in the native. The radius of gyration of the acid-induced folded state at pH 0 is shown to be identical to that of the native state at pH 7 by small angle X-ray scattering. We conclude that acid-induced collapse of Sac7d does not lead to a molten globule but proceeds directly to the native state. The folding of Sac7d as a function of pH and anion concentration is summarized with a phase diagram that is similar to those observed for other proteins that undergo acid-induced folding except that the A-state is encompassed by the native state. These results demonstrate that formation of a molten globule is not a general property of proteins that are refolded by acid. [source]


Interactions of trifluroethanol with the Trp-cage peptide

BIOPOLYMERS, Issue 2-3 2007
Chiradip Chatterjee
Abstract It has been suggested that aggregation of fluorinated alcohols in water solutions is involved with the abilities of these alcohols to provoke conformational changes in peptides and proteins. The extent of fluoroalcohol aggregation depends on the degree of fluorination: hexafluoroisopropanol (HFIP) is more extensively aggregated than is TFE. We previously described a study of the interactions of HFIP with the peptide Trp-cage and provided evidence for the formation of long-lived complexes between this fluoroalcohol and the peptide. In the present work, we have examined the interactions of the less-fluorinated TFE with Trp-cage, in order to probe the role of fluoroalcohol aggregation in the phenomena observed. Intermolecular 1H{19F} nuclear Overhauser effects arising from interactions of TFE with the hydrogens of the peptide in a solution containing 42% TFE were determined at sample temperatures from 5 to 45°C. It is shown that the folded state of the peptide under these conditions is essentially the same as that observed in water and in 30% HFIP-water. The observed peptide,solvent NOEs indicate formation of complexes of Trp-cage with TFE that persist for times of the order of 1 ns. The interactions leading to complexes with TFE are somewhat weaker than those involved in complex formation with HFIP. There are no indications that the aggregation of fluoroalcohol is a necessary concomitant of the interactions of TFE or HFIP with Trp-cage. Rather, the stronger and more long-lived interactions of HFIP with Trp-cage appear to be primarily the result of the greater hydrogen-bonding ability and hydrophobicity of this fluoroalcohol. © 2007 Wiley Periodicals, Inc. Biopolymers 87: 115,123, 2007. This article was originally published online as an accepted preprint. The "Published Online" date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com [source]


Does the anesthetic 2,2,2-trifluoroethanol interact with bovine serum albumin by direct binding or by solvent-mediated effects?

BIOPOLYMERS, Issue 2 2005
A calorimetric, spectroscopic investigation
Abstract Thermal unfolding of bovine serum albumin (BSA) has been studied in the presence of 2,2,2-trifluoroethanol (TFE) using high-sensitivity microdifferential scanning calorimetry. Quantitative thermodynamic parameters accompanying the thermal transitions have been evaluated. TFE is observed to be a stabilizer or a destabilizer of the folded state of BSA depending on the pH. CD spectroscopy revealed an increase in the ,-helical content of BSA and a decrease in the tertiary structure in the presence of increasing molalities of TFE. Isothermal titration calorimetric results do not indicate appreciable binding of the TFE molecules to BSA. TFE quenches the steady-state tryptophan fluorescence of BSA only at higher molalities and there is no effect on the tryptophan fluorescence at lower molalities. The calorimetric and spectroscopic results obtained in this work suggest that solvent-mediated effects dominate the interaction of TFE with BSA and the binding component may be very weak. Since the binding component is very weak, one of the possibilities of anesthetic action of TFE molecules on the actual targets may be through perturbation of the structural and dynamic properties of the lipid bilayer so that the function of crucial but unspecified membrane proteins is affected. © 2005 Wiley Periodicals, Inc. Biopolymers 78: 78,86, 2005 [source]


Structure and lability of archaeal dehydroquinase

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 10 2008
Natasha N. Smith
Multiple sequence alignments of type I 3-dehydroquinate dehydratases (DQs; EC 4.2.1.10) show that archaeal DQs have shorter helical regions than bacterial orthologs of known structure. To investigate this feature and its relation to thermostability, the structure of the Archaeoglobus fulgidus (Af) DQ dimer was determined at 2.33,Ć resolution and its denaturation temperature was measured in vitro by circular dichroism (CD) and differential scanning calorimetry (DSC). This structure, a P212121 crystal form with two 45,kDa dimers in the asymmetric unit, is the first structural representative of an archaeal DQ. Denaturation occurs at 343 ± 3,K at both low and high ionic strength and at 349,K in the presence of the substrate analog tartrate. Since the growth optimum of the organism is 356,K, this implies that the protein maintains its folded state through the participation of additional factors in vivo. The (,,)8 fold is compared with those of two previously determined type I DQ structures, both bacterial (Salmonella and Staphylococcus), which had sequence identities of ,30% with AfDQ. Although the overall folds are the same, there are many differences in secondary structure and ionic features; the archaeal protein has over twice as many salt links per residue. The archaeal DQ is smaller than its bacterial counterparts and lower in regular secondary structure, with its eight helices being an average of one turn shorter. In particular, two of the eight normally helical regions (the exterior of the barrel) are mostly nonhelical in AfDQ, each having only a single turn of 310 -helix flanked by ,-strand and coil. These `protohelices' are unique among evolutionarily close members of the (,,)8 -fold superfamily. Structural features that may contribute to stability, in particular ionic factors, are examined and the implications of having a Tm below the organism's growth temperature are considered. [source]