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

The goal of this article is to gain a better understanding of the denatured state ensemble (DSE) of proteins through an experimental and computational study of their denaturation by urea. Proteins unfold to different extents in urea and the most hydrophobic proteins have the most compact DSE and contain almost as much secondary structure as folded proteins. Proteins that unfold to the greatest extent near pH 7 still contain substantial amounts of secondary structure.

NMR study and molecular dynamics simulations of optimized beta-hairpin fragments of protein G.

The stability and structure of several beta-hairpin peptide variants derived from the C-terminus of the B1 domain of protein G were investigated by a number of experimental and computational techniques. Our analysis shows that the structure and stability of this hairpin can be greatly affected by one or a few simple mutations. For example, removing an unfavorable charge near the N-terminus of the peptide (Glu42 to Gln or Thr) or optimization of the N-terminal charge-charge interactions (Gly41 to Lys) both stabilize the peptide, even in water.

Terminal ion pairs stabilize the second beta-hairpin of the B1 domain of protein G.

The effects of terminal ion pairs on the stability of a beta-hairpin peptide corresponding to the C-terminal residues of the B1 domain of protein G were determined using thermal unfolding as monitored by nuclear magnetic resonance and circular dichroism spectroscopy. Molecular dynamics (MD) simulations were also performed to examine the effect of ion pairs on the structures. Eight peptides were studied including the wild type (G41) and the N-terminal modified sequences that had the first residue deleted (E42), replaced with a Lys (K41), or extended by an additional Gly (G40).

pK values of histidine residues in ribonuclease Sa: effect of salt and net charge.

The primary goal of this study was to gain a better understanding of the effect of environment and ionic strength on the pK values of histidine residues in proteins. The salt-dependence of pK values for two histidine residues in ribonuclease Sa (RNase Sa) (pI=3.5) and a variant in which five acidic amino acids have been changed to lysine (5K) (pI=10.

Charge-charge interactions are key determinants of the pK values of ionizable groups in ribonuclease Sa (pI=3.5) and a basic variant (pI=10.2).

The pK values of the titratable groups in ribonuclease Sa (RNase Sa) (pI=3.5), and a charge-reversed variant with five carboxyl to lysine substitutions, 5K RNase Sa (pI=10.2), have been determined by NMR at 20 degrees C in 0.

Charge-charge interactions are the primary determinants of the pK values of the ionizable groups in Ribonuclease T1.

Coulomb's law and a finite difference Poisson-Boltzmann based analysis are used to predict the pK values for 15 ionizable side chains (6 Asp, 6 Glu and 3 His) in ribonuclease T1. These predicted values are compared to the measured pK values to gain insight into the most important factors that influence the pK values of the ionizable groups in proteins. Charge-charge interactions are clearly the most important factor that determines the pK values of most ionizable groups in ribonuclease T1.