Determining stability of proteins from guanidinium chloride transition curves.
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ABSTRACT:
SUBMITTER: Ahmad F
PROVIDER: S-EPMC1133190 | biostudies-other | 1992 Oct
REPOSITORIES: biostudies-other
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Project description:The methionine adenosyltransferase from the thermophile Methanococcus jannaschii is fully and irreversibly unfolded in the presence of guanidinium chloride. Unfolding of this dimeric protein is a three-state process in which a dimeric intermediate could be identified. The less stable secondary structural elements of the protein are the C-terminal ends of ?-strands E2 and E6, as deduced from the behavior of tyrosine to tryptophan mutants at residues 72 and 170, which are located in the subunit interface. Unraveling of these elements at the monomer interface may soften intersubunit interactions, leading to the observed 85% activity loss. Accumulation of the intermediate was associated with maintenance of residual activity, an increase in the elution volume of the protein upon gel filtration and a decrease in the sedimentation coefficient. Elimination of the remaining enzymatic activity occurred in conjunction with a 50% reduction in helicity and fluorescence alterations illustrating a transient burial of tryptophans at ?-strands E2, E3 and E9. The available 3D-model predicted that these ?-strands are involved in the central and N-terminal domains of the monomer structure. Severe perturbation of this area of the monomer-monomer interface may destroy the remaining intermolecular interactions, thus leading to dissociation and aggregation. Finally, transition to the denatured state includes completion of the changes detected in the microenvironments around tryptophans included at ?-helixes H5 and H6, the loops connecting H5-E8 and E9, ?-strands E3 and E12.
| S-EPMC3090699 | biostudies-literature
Project description:The denatured states of proteins have always attracted our attention due to the fact that the denatured state is the only experimentally achievable state of a protein, which can be taken as initial reference state for considering the in vitro folding and defining the native protein stability. It is known that heat and guanidinium chloride (GdmCl) give structurally different states of RNase-A, lysozyme, ?-chymotrypsinogen A and ?-lactalbumin. On the contrary, differential scanning calorimetric (DSC) and isothermal titration calorimetric measurements, reported in the literature, led to the conclusion that heat denatured and GdmCl denatured states are thermodynamically and structurally identical. In order to resolve this controversy, we have measured changes in the far-UV CD (circular dichroism) of these heat-denatured proteins on the addition of different concentrations of GdmCl. The observed sigmoidal curve of each protein was analyzed for Gibbs free energy change in the absence of the denaturant (?G0X?D) associated with the process heat denatured (X) state ? GdmCl denatured (D) state. To confirm that this thermodynamic property represents the property of the protein alone and is not a manifestation of salvation effect, we measured urea-induced denaturation curves of these heat denatured proteins under the same experimental condition in which GdmCl-induced denaturation was carried out. In this paper we report that (a) heat denatured proteins contain secondary structure, and GdmCl (or urea) induces a cooperative transition between X and D states, (b) for each protein at a given pH and temperature, thermodynamic cycle connects quantities, ?G0N?X (native (N) state ? X state), ?G0X?D and ?G0N?D (N state ? D state), and
| S-EPMC4457810 | biostudies-literature
Project description:In the title compound, C(20)H(18)N(3)O(+)·Cl(-), the orientation of the aromatic rings around the planar CN(3) (+) unit produces steric hindrance. As a consequence of this particular orientation of the guanidinium cation, hydrogen bonding is restricted to N-H?Cl and intra-molecular N-H?O hydrogen bonds within the discrete unit. The guanidinium and carbonyl groups are coplanar as a result of the six-membered ring formed by the N-H?O intra-molecular hydrogen bond. The dihedral angles between the guanidinium plane and the two phenyl rings are 62.31?(8) and 64.24?(8)°.
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Project description:The non-H atoms of the cation of the title salt, C(8)H(9)N(4)O(+)·Cl(-), are approximately co-planar (r.m.s. deviation = 0.024?Å) with one amino group forming an intra-molecular hydrogen bond to the tertiary N atom of the benzoxazole fused-ring system. The cations and anions are linked by cyclic R(2) (1)(6) N-H?Cl hydrogen-bonding associations, generating linear chains running along the a-axis direction.
| S-EPMC3247517 | biostudies-literature
Project description:In the crystal of the title compound, 2CH(6)N(3) (+)·2Cl(-)·C(12)H(24)O(6), the 18-crown-6 mol-ecule is located across an inversion center. The guanidinium cation links to the 18-crown-6 mol-ecule and chloride anion via N-H?O and N-H?Cl hydrogen bonds.
| S-EPMC3344600 | biostudies-literature
Project description:1. Caesium chloride and guanidinium chloride were shown to cause conformational changes in the high-molecular-weight mucoprotein A of water-soluble gastric mucus with no change in molecular weight. 2. Increasing concentrations of CsCl decrease the viscosity of the mucoprotein bringing about a transition which is essentially complete in 0.1m-CsCl. The shear-dependence of viscosity of the mucoprotein is abolished by low concentrations of CsCl. The normally highly expanded molecule becomes contracted in CsCl to a molecule having the same symmetry but a smaller volume and decreased solvation, in keeping with an increased sedimentation coefficient (18.7S-->33S). 3. This contracted form does not revert to the native conformation on removal of the CsCl. 4. A mechanism is discussed in terms of the effect of the Cs(+) and Cl(-)ions on water structure and the water-mucoprotein interaction. 5. Guanidinium chloride causes the CsCl-treated material to expand, in keeping with a decrease in s(0) (25,w) (33S-->26S). This is analogous to the known unfolding effect of guanidinium chloride on proteins and suggests that guanidinium chloride solubilizes groups involved in stabilizing the contracted structure. Removal of the guanidinium chloride results in a limited aggregation of four mucoprotein molecules. 6. These results show that caution must be exercised before interpreting the physical properties of mucoproteins which have been treated with CsCl and/or guanidinium chloride.
| S-EPMC1168168 | biostudies-other
Project description:The non-H atoms of the cation of the title salt, C(8)H(9)N(4)S(+)·Cl(-), are approximately co-planar (r.m.s. deviation = 0.037?Å), with one amino group forming an intra-molecular hydrogen bond to the tertiary N atom of the benzothia-zole fused-ring system. The cations and anions are linked by cyclic R(2) (1)(6) N-H?Cl hydrogen-bonding associations, generating helical chains running along the b-axis direction.
| S-EPMC3247516 | biostudies-other
Project description:The guanidinium cation (C(NH2)3(+)) is a highly stable cation in aqueous solution due to its efficient solvation by water molecules and resonance stabilization of the charge. Its salts increase the solubility of nonpolar molecules ("salting-in") and decrease the ordering of water. It is one of the strongest denaturants used in biophysical studies of protein folding. We investigate the behavior of guanidinium and its derivative, methyl guanidinium (an amino acid analogue) at the air-water surface, using atomistic molecular dynamics (MD) simulations and calculation of potentials of mean force. Methyl guanidinium cation is less excluded from the air-water surface than guanidinium cation, but both cations show orientational dependence of surface affinity. Parallel orientations of the guanidinium ring (relative to the Gibbs dividing surface) show pronounced free energy minima in the interfacial region, while ring orientations perpendicular to the GDS exhibit no discernible surface stability. Calculations of surface fluctuations demonstrate that, near the air-water surface, the parallel-oriented cations generate significantly greater interfacial fluctuations compared to other orientations, which induces more long-ranged perturbations and solvent density redistribution. Our results suggest a strong correlation with induced interfacial fluctuations and ion surface stability. These results have implications for interpreting molecular-level, mechanistic action of this osmolyte's interaction with hydrophobic interfaces as they impact protein denaturation (solubilization).
| S-EPMC4216681 | biostudies-literature
Project description:The conformational stability of the ?-hairpin miniprotein, CLN025, a variant of chignolin in which the N- and C-terminal glycines are replaced by tyrosines, in various concentrations of guanidinium chloride (GdmCl) and urea was examined by molecular dynamics simulations and electronic circular dichroism (ECD) spectropolarimetry. The peptide maintains its ?-hairpin conformation in GdmCl and urea solutions. In GdmCl, Gly7 influences the turn to reduce the number of Asp3-Gly7 H-bonds and the Tyr1-Trp9 H-bond is lost. The structure of the peptide is less stable in 3 M GdmCl than in water or 6 M GdmCl, because the number of Asp3-Thr8 and Tyr1-Tyr10 H-bonds are reduced and the Tyr2 side chain moves away from the Pro4 and Trp9 side chains and toward the Tyr10 side chain. This reduces the number of Tyr2-Pro4 CH-? interactions and Tyr2-Trp9 and Tyr1-Tyr10 aromatic-aromatic (Ar-Ar) interactions and increases the number of Tyr2-Tyr10 Ar-Ar interactions. In 6 M GdmCl at 300 and 333 K, the number of Tyr1-Tyr10 and Asp3-Thr8 H-bonds increases, but fewer structures have Tyr2-Pro4 CH-? and Tyr1-Tyr10 and Tyr2-Trp9 Ar-Ar interactions. In urea, Gly7 is in a mixture of ?-turn and random meander structures and the number of Asp3-Thr6 and Tyr1-Tyr10 H-bonds are reduced as are the number of Tyr2-Pro4 CH-? interactions and Tyr1-Tyr10 and Tyr2-Trp9 Ar-Ar interactions. In 4 M urea, a shorter turn places Gly7 into the ?-sheet region and Tyr10 is pushed out into the solvent. In 8 M urea, the number of Asp3-Glu5 H-bonds is increased and the ?-sheet is lost, but the electrostatic interaction between the charged termini is restored and a cation-? interaction between the indolyl ring of Trp9 and the positively charged N-terminus is formed. In 8 M urea at 333 K, the ?-hairpin conformation is almost lost. The structure of CLN025 is stable, because the weakly polar interactions and H-bonds maintain the ?-hairpin conformation in the various environments. CLN025 should not be considered a miniprotein, because it lacks a well-defined tertiary structure, it is resistant to denaturation, it does not have an increased heat capacity near its melting temperature, and the structures near and above the melting temperature retain a ?-hairpin conformation.
| S-EPMC3096019 | biostudies-literature
Project description:Avidin molecules in which a fraction of the four binding sites were occupied by biotin did not dissociate completely in 6.4m-guanidinium chloride. Only unoccupied subunits dissociated. The remainder recombined to form the tetrameric avidin-biotin complex. The rate at which unoccupied subunits were unfolded and dissociated was only decreased by one-half in species in which three of the four binding sites were occupied by biotin. These results can be explained by assuming that unfolding of unoccupied subunits followed by dissociation from the tetramer is initiated by penetration of guanidinium ions into the binding site and disorganization of this region of the subunit. When a site is occupied by biotin this pathway is blocked and the subunit does not unfold. Each subunit behaves independently and is not markedly stabilized when neighbouring subunits are occupied.
| S-EPMC1174509 | biostudies-other