Relationship of Leffler (Brønsted) α values and protein folding Φ values to position of transition-state structures on reaction coordinates

Self Cite

The presence of a solvent-exposed alanine residue stabilizes a helix by 0.4 -2 kcal⅐mol ؊1 relative to glycine. Various factors have been suggested to account for the differences in helical propensity, from the higher conformational freedom of glycine sequences in the unfolded state to hydrophobic and van der Waals' stabilization of the alanine side chain in the helical state. We have performed all-atom molecular dynamics simulations with explicit solvent and exhaustive sampling of model peptides to address the backbone conformational entropy difference between Ala and Gly in the denatured state. The mutation of Ala to Gly leads to an increase in conformational entropy equivalent to Ϸ0.4 kcal⅐mol ؊1 in a fully flexible denatured, that is, unfolded, state. But, this energy is closely counterbalanced by the (measured) difference in free energy of transfer of the glycine and alanine side chains from the vapor phase to water so that the unfolded alanineand glycine-containing peptides are approximately isoenergetic. The helix-stabilizing propensity of Ala relative to Gly thus mainly results from more favorable interactions of Ala in the folded helical structure. The small difference in energetics in the denatured states means that the ⌽-values derived from Ala 3 Gly scanning of helices are a very good measure of the extent of formation of structure in proteins with little residual structure in the denatured state.folding ͉ pathway ͉ protein ͉ stability ͉ transition state

Section: Discussionmentioning

confidence: 99%

Conformational entropy of alanine versus glycine in protein denatured states

Scott

Alonso

Sato

et al. 2007

Self Cite

“…As an example, in the I 3 V 3 A 3 G mutation series. the I 3 V measures interactions originating from tertiary structure contacts, the V 3 A measures a mixture of tertiary and secondary structure interactions, whereas the A 3 G reports almost exclusively on secondary structure formation (33).…”

Section: In This Context Deviations From Linearity May Indicate Ts Smentioning

confidence: 99%

“…In multipoint plots, different local probes of the same residue are forced in a single fit that can yield wrong estimates (33). As an example, in the I 3 V 3 A 3 G mutation series.…”

Section: In This Context Deviations From Linearity May Indicate Ts Smentioning

confidence: 99%

Φ-Value analysis by molecular dynamics simulations of reversible folding

Settanni

Rao

Caflisch

2005

In ⌽-value analysis, the effects of mutations on the folding kinetics are compared with the corresponding effects on thermodynamic stability to investigate the structure of the protein-folding transition state (TS). Here, molecular dynamics (MD) simulations (totaling 0.65 ms) have been performed for a large set of single-point mutants of a 20-residue three-stranded antiparallel ␤-sheet peptide. Between 57 and 120 folding events were sampled at near equilibrium for each mutant, allowing for accurate estimates of folding͞unfolding rates and stability changes. The ⌽ values calculated from folding and unfolding rates extracted from the MD trajectories are reliable if the stability loss upon mutation is larger than Ϸ0.6 kcal͞mol, which is observed for 8 of the 32 single-point mutants. The same heterogeneity of the TS of the wild type was found in the mutated peptides, showing two possible pathways for folding. Single-point mutations can induce significant TS shifts not always detected by ⌽-value analysis. Specific nonnative interactions at the TS were observed in most of the peptides studied here. The interpretation of ⌽ values based on the ratio of atomic contacts at the TS over the native state, which has been used in the past in MD and Monte Carlo simulations, is in agreement with the TS structures of wild-type peptide. However, ⌽ values tend to overestimate the nativeness of the TS ensemble, when interpreted neglecting the nonnative interactions. ⌽ values are usually interpreted in terms of native contacts (4). This description has been successfully used to obtain sets of conformations from the TS ensemble of several proteins (5-9) and to bias molecular dynamics (MD) trajectories toward the TS (10). On the other hand, specific nonnative interactions may be formed at both the TS and denatured-state ensemble and lead to a wrong picture of TS if not taken into account (11). Furthermore, different experimental conditions or mutations may determine detectable changes in the TS structure, showing the presence of parallel pathways (12, 13) and, thus, a heterogeneous TS. In addition, the ensemble average associated with the use of certain folding observables, like the degree of tryptophan burial, may disguise the presence of multiple folding pathways and folding intermediates (14). Namely, a recent study (15) suggests that not all conformations obtained in MD simulations by using ⌽ values as restraints on a subset of the native contacts belong to the TS.The TS structures can be identified by MD simulations through the calculation of their folding probability P fold (16), i.e., the probability that a trajectory started from a given structure reaches the folded state before unfolding. The concept of P fold calculation was first introduced in a method for determining transmission coefficients, starting from a known TS (17), and used to identify TSs of simple conformational changes (e.g., tyrosine ring flips) (18). The approach has recently been used to study the otherwise very elusive folding TS by atomistic Monte Carlo off...

“…The slope of the observed correlation, classically denoted as α, reflects the position of the transition state along the reaction coordinate. Surprisingly, LFER plots not only apply to simple organic reactions but also to complex reactions stabilized by many noncovalent interactions, and have been then used in enzymology (25), binding reactions involving allosteric control (26,27), and protein folding (28).…”

Section: An Ordered Transition State As a Signature Of Geometrical Prmentioning

confidence: 99%

Structure of the transition state for the binding of c-Myb and KIX highlights an unexpected order for a disordered system

Giri

Morrone

Toto

et al. 2013

100

133

A classical dogma of molecular biology dictates that the 3D structure of a protein is necessary for its function. However, a considerable fraction of the human proteome, although functional, does not adopt a defined folded state under physiological conditions. These intrinsically disordered proteins tend to fold upon binding to their partners with a molecular mechanism that is elusive to experimental characterization. Indeed, although many hypotheses have been put forward, the functional role (if any) of disorder in these intrinsically denatured systems is still shrouded in mystery. Here, we characterize the structure of the transition state of the binding-induced folding in the reaction between the KIX domain of the CREB-binding protein and the transactivation domain of c-Myb. The analysis, based on the characterization of a series of conservative site-directed mutants, reveals a very high content of native-like structure in the transition state and indicates that the recognition between KIX and c-Myb is geometrically precise. The implications of our results in the light of previous work on intrinsically unstructured systems are discussed.kinetics | mutagenesis | protein folding