In Φ-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 β-sheet peptide. Between 57 and 120 folding events were sampled at near equilibrium for each mutant, allowing for accurate estimates of folding/unfolding rates and stability changes. The Φ values calculated from folding and unfolding rates extracted from the MD trajectories are reliable if the stability loss upon mutation is larger than approximately 0.6 kcal/mol, 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 Φ-value analysis. Specific nonnative interactions at the TS were observed in most of the peptides studied here. The interpretation of Φ 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, Φ values tend to overestimate the nativeness of the TS ensemble, when interpreted neglecting the nonnative interactions.
BT - Proceedings of the National Academy of Sciences of the United States of America C1 - 0064 DA - 2005 Jan 18 DO - 10.1073/pnas.0406754102 IS - 3 J2 - Proc. Natl. Acad. Sci. U.S.A. LA - eng M3 - Research Article N2 -In Φ-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 β-sheet peptide. Between 57 and 120 folding events were sampled at near equilibrium for each mutant, allowing for accurate estimates of folding/unfolding rates and stability changes. The Φ values calculated from folding and unfolding rates extracted from the MD trajectories are reliable if the stability loss upon mutation is larger than approximately 0.6 kcal/mol, 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 Φ-value analysis. Specific nonnative interactions at the TS were observed in most of the peptides studied here. The interpretation of Φ 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, Φ values tend to overestimate the nativeness of the TS ensemble, when interpreted neglecting the nonnative interactions.
PY - 2005 SP - 628 EP - 633 T2 - Proceedings of the National Academy of Sciences of the United States of America TI - Phi-value analysis by molecular dynamics simulations of reversible folding VL - 102 SN - 0027-8424 ER -