Urea denatured state ensembles contain extensive secondary structure that is increased in hydrophobic proteins

Pace, C. Nick; Huyghues-Despointes, Beatrice M. P.; Fu, Hailong; Takano, Kazufumi; Scholtz, J. Martin; Grimsley, Gerald R.

doi:10.1002/pro.370

Cited by 40 publications

(14 citation statements)

References 102 publications

(127 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, the principal cause for the scatter of the data around the identity line is likely to be the denatured state structure. It is known that the compactness of the denatured state depends both on the osmolyte used [12, 40], and on the type of protein [41]. …”

Section: Resultsmentioning

confidence: 99%

Osmolyte effects on protein stability and solubility: A balancing act between backbone and side-chains

Auton

Rösgen

Sinev

et al. 2011

Biophysical Chemistry

228

261

View full text Add to dashboard Cite

In adaptation biology the discovery of intracellular osmolyte molecules that in some cases reach molar levels, raises questions of how they influence protein thermodynamics. We’ve addressed such questions using the premise that from atomic coordinates, the transfer free energy of a native protein (ΔGtr,N) can be predicted by summing measured water-to-osmolyte transfer free energies of the protein’s solvent exposed side chain and backbone component parts. ΔGtr,D is predicted using a self avoiding random coil model for the protein, and ΔGtr,D − ΔGtr,N, predicts the m-value, a quantity that measures the osmolyte effect on the N ⇌ D transition. Using literature and newly measured m-values we show 1:1 correspondence between predicted and measured m-values covering a range of 12 kcal/mol/M in protein stability for 46 proteins and 9 different osmolytes. Osmolytes present a range of side chain and backbone effects on N and D solubility and protein stability key to their biological roles.

show abstract

Section: Resultsmentioning

confidence: 99%

Osmolyte effects on protein stability and solubility: A balancing act between backbone and side-chains

Auton

Rösgen

Sinev

et al. 2011

Biophysical Chemistry

228

261

View full text Add to dashboard Cite

show abstract

“…More likely, the correct extent of exposure is intermediate between the upper and lower bound models. 62, 63 …”

Section: Discussionmentioning

confidence: 99%

“…63, 94 RNases Sa, T1, and their variants were expressed and purified as described previously. 82, 95, 96 The purity of all proteins was confirmed by SDS PAGE and MALDI-TOF mass spectrometry.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Contribution of Hydrophobic Interactions to Protein Stability

Pace

Fryar

et al. 2011

Journal of Molecular Biology

Self Cite

351

229

View full text Add to dashboard Cite

Our goal was to gain a better understanding of the contribution of hydrophobic interactions to protein stability. We measured the change in conformational stability, Δ(ΔG), for hydrophobic mutants of four proteins: villin head piece subdomain (VHP) with 36 residues, a surface protein from Borrelia burgdorferi (VlsE) with 341 residues, and two proteins previously studied in our laboratory, ribonucleases Sa and T1. We compare our results with previous studies and reach the following conclusions. 1. Hydrophobic interactions contribute less to the stability of a small protein, VHP (0.6 ± 0.3 kcal/mole per –CH2– group), than to the stability of a large protein, VlsE (1.6 ± 0.3 kcal/mol per –CH2– group). 2. Hydrophobic interactions make the major contribution to the stability of VHP (40 kcal/mol) and the major contributors are (in kcal/mol): Phe 18 (3.9), Met 13 (3.1), Phe 7 (2.9), Phe 11 (2.7), and Leu 21 (2.7). 3. Based on Δ(ΔG) values for 148 hydrophobic mutants in 13 proteins, burying a –CH2– group on folding contributes, on average, 1.1 ± 0.5 kcal/mol to protein stability. 4. The experimental Δ(ΔG) values for aliphatic side chains (Ala, Val, Ile, and Leu) are in good agreement with their ΔGtr values from water to cyclohexane. 5. For 22 proteins with 36 to 534 residues, hydrophobic interactions contribute 60 ± 4% and hydrogen bonds 40 ± 4% to protein stability. 6. Conformational entropy contributes about 2.4 kcal/mol per residue to protein instability. The globular conformation of proteins is stabilized predominately by hydrophobic interactions.

show abstract

“…MD studies have shown that chignolin is also resistant to unfolding, even at 373 K. 45 Proteins with compact denatured state ensembles contain a greater secondary structure content 46, 47 that reduces the number of interactions between protein and denaturant. 48 CLN025 is a stable β-hairpin polypeptide that is composed entirely of secondary structure with no tertiary structure. Therefore, it is not affected by denaturant and does not unfold as would a protein with tertiary structure.…”

Section: Cln025 Is a Stable Polypeptide Model Of Secondary Structurementioning

confidence: 99%

The CLN025 Decapeptide Retains a β-Hairpin Conformation in Urea and Guanidinium Chloride

2011

View full text Add to dashboard Cite

The conformational stability of the β-hairpin miniprotein, CLN025, a variant of chignolin in which the N- and C-terminal Gly are replaced by Tyr, in various concentrations of GdmCl and urea was examined by Molecular Dynamics simulations and Electronic Circular Dichroism (ECD) spectropolarimetry. The peptide maintains its β-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 towards the Tyr10 side-chain. This reduces the number of Tyr2-Pro4 CH-π 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-π and Tyr1-Tyr10 and Tyr2-Trp9 Ar-Ar interactions. In urea, Gly7 is in a mixture of β-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-π interactions and Tyr1-Tyr10 and Tyr2-Trp9 Ar-Ar interactions. In 4 M urea, a shorter turn places Gly7 in to the β-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 β-sheet is lost, but the electrostatic interaction between the charged termini is restored and a cation-π interaction between the indolyl ring of Trp9 and the positively charged N-terminus is formed. In 8 M urea at 333 K, the β-hairpin conformation is almost lost. The structure of CLN025 is stable, because the weakly polar interactions and H-bonds maintain the β-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, does not have an increased heat capacity near its melting temperature and the structures near and above the melting temperature retain a β-hairpin conformation.

show abstract

Urea denatured state ensembles contain extensive secondary structure that is increased in hydrophobic proteins

Cited by 40 publications

References 102 publications

Osmolyte effects on protein stability and solubility: A balancing act between backbone and side-chains

Osmolyte effects on protein stability and solubility: A balancing act between backbone and side-chains

Contribution of Hydrophobic Interactions to Protein Stability

The CLN025 Decapeptide Retains a β-Hairpin Conformation in Urea and Guanidinium Chloride

Contact Info

Product

Resources

About