Charge–charge interactions in the denatured state influence the folding kinetics of ribonuclease Sa

Trefethen, Jared M.; Pace, C. Nick; Scholtz, J. Martin; Brems, David N.

doi:10.1110/ps.051401905

Cited by 46 publications

(48 citation statements)

References 23 publications

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“…Here DSE effects are important and the small φ-value indicates just the opposite, namely that the non-native interaction probed by the mutation is present in the transition state but is slightly weakened relative to the DSE. Similar effects have been observed by the Texas A&M group with their studies of the role of surface acidic residues in the folding ribonuclease Sa [30].…”

Section: The Role Of Non-native Dse Electrostatic Interactions In Folsupporting

confidence: 82%

“…This is an important observation since it shows that DSE electrostatic interactions can make large contributions to protein stability. Significant DSE electrostatic interactions have also been observed in RNase H and ribonuclease Sa [29][30][31]. The case of RNase H is particularly interesting because it provides an example of the possible functional consequences of DSE structure.…”

Section: Electrostatic Interactions Are Found In the Denatured State mentioning

confidence: 99%

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Electrostatic interactions in the denatured state ensemble: Their effect upon protein folding and protein stability

Cho¹,

Sato²,

Horng³

et al. 2008

Archives of Biochemistry and Biophysics

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It is now recognized that the denatured state ensemble (DSE) of proteins can contain significant amounts of structure, particularly under native conditions. Well-studied examples include small units of hydrogen bonded secondary structure, particularly helices or turns as well hydrophobic clusters. Other types of interactions are less well characterized and it has often been assumed that electrostatic interactions play at most a minor role in the DSE. However, recent studies have shown that both favorable and unfavorable electrostatic interactions can be formed in the DSE. These can include surprisingly specific non-native interactions that can even persist in the transition state for protein folding. DSE electrostatic interactions can be energetically significant and their modulation either by mutation or by varying solution conditions can have a major impact upon protein stability. pH dependent stability studies have shown that electrostatic interactions can contribute up to 4 kcal mol −1 to the stability of the DSE.

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Section: The Role Of Non-native Dse Electrostatic Interactions In Folsupporting

confidence: 82%

Section: Electrostatic Interactions Are Found In the Denatured State mentioning

confidence: 99%

Electrostatic interactions in the denatured state ensemble: Their effect upon protein folding and protein stability

Cho¹,

Sato²,

Horng³

et al. 2008

Archives of Biochemistry and Biophysics

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“…Mutations that alter the energetics of the DSE can impact the analysis of cooperativity and folding, and could modulate the propensity to aggregate. other proteins, thus NTL9 is not an isolated example (4,10,14,18,22,27).…”

Section: Resultsmentioning

confidence: 99%

“…There is growing evidence that the DSE can be compact under native conditions and can form native and nonnative clusters of hydrophobic residues, native and nonnative secondary structure, and even nonnative electrostatic interactions, thus the NTL9 is unlikely to be a special case (10,12,14,18,22,27,46,47). NTL9 is emerging as a popular system for long molecular dynamics simulations and the data presented here will help provide a rigorous benchmark for these efforts (45,(48)(49)(50).…”

Section: Discussionmentioning

confidence: 97%

Energetically significant networks of coupled interactions within an unfolded protein

Cho

Meng

Sato

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Unfolded and partially unfolded proteins participate in a wide range of biological processes from pathological aggregation to the regulation of normal cellular activity. Unfolded states can be populated under strongly denaturing conditions, but the ensemble which is relevant for folding, stability, and aggregation is that populated under physiological conditions. Characterization of nonnative states is critical for the understanding of these processes, yet comparatively little is known about their energetics and their structural propensities under native conditions. The standard view is that energetically significant coupled interactions involving multiple residues are generally not present in the denatured state ensemble (DSE) or in intrinsically disordered proteins. Using the N-terminal domain of the ribosomal protein L9, a small α-β protein, as an experimental model system, we demonstrate that networks of energetically significant, coupled interactions can form in the DSE of globular proteins, and can involve residues that are distant in sequence and spatially well separated in the native structure. X-ray crystallography, NMR, dynamics studies, native state pK a measurements, and thermodynamic analysis of more than 25 mutants demonstrate that residues are energetically coupled in the DSE. Altering these interactions by mutation affects the stability of the domain. Mutations that alter the energetics of the DSE can impact the analysis of cooperativity and folding, and may play a role in determining the propensity to aggregate.protein folding | protein stability | ϕ-value analysis | double-mutant cycle T he properties of nonnative states of proteins have attracted considerable attention because they can play critical roles in cell signaling, translocation across membranes, temperaturesensitive phenotypes, and in a wide range of protein deposition diseases (1-14). The properties of unfolded states are also relevant to any discussion of intrinsically disordered proteins (IDPs) (1). The denatured state ensemble (DSE) can be populated under strongly denaturing conditions and studied at equilibrium; however, the physiologically relevant DSE is the state that is in equilibrium with the folded state under native conditions. This state is difficult to study directly and most studies of the DSE under native conditions have made use of indirect approaches. Nevertheless, experimental and theoretical work has shown that the DSE populated under physiological conditions is not a random coil, but can contain native and nonnative structure, stabilized by local and nonlocal interactions. However, very little is known about the energetics of these interactions, particularly ones involving residues distant in sequence, or about energetically coupled interactions involving multiple residues in the . These are a critical piece of the folding puzzle; they impact protein aggregation and folding, and have important implications for the properties of IDPs. Recent studies offer evidence for hydrophobic clusters, electrostatic interactio...

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“…There is now considerable evidence showing that electrostatic interactions in the DSE can affect both the stability and kinetics of folding of proteins. 75,76 Another observation is interesting. It is clear that an excess of positive charges on the molecules causes a much larger increase in the m value than an excess of negative charges.…”

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confidence: 92%

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

Pace

Huyghues-Despointes

et al. 2010

Protein Science

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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. At low pH, the DSE expands due to charge-charge interactions and when the net charge per residue is high, most of the secondary structure is disrupted. The proteins in the DSE appear to contain substantial amounts of polyproline II conformation at high urea concentrations. In all cases considered, including staph nuclease, the extent of unfolding by urea can be accounted for using the data and approach developed in the laboratory of Wayne Bolen (Auton et al., Proc Natl Acad Sci 2007; 104:15317-15323).

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Charge–charge interactions in the denatured state influence the folding kinetics of ribonuclease Sa

Cited by 46 publications

References 23 publications

Electrostatic interactions in the denatured state ensemble: Their effect upon protein folding and protein stability

Electrostatic interactions in the denatured state ensemble: Their effect upon protein folding and protein stability

Energetically significant networks of coupled interactions within an unfolded protein

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

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