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The mechanism of folding of Im7 reveals competition between functional and kinetic evolutionary constraints.

Friel CT, Smith DA, Vendruscolo M, Gsponer J, Radford SE - Nat. Struct. Mol. Biol. (2009)

Bottom Line: It is not clear whether this type of folding landscape results from insufficient evolutionary pressure to optimize folding efficiency, or arises from a conflict between functional and folding constraints.The results provide a comprehensive view of the folding process of this small protein.An analysis of the contributions of native and non-native interactions at different stages of folding reveals how the complexity of the folding landscape arises from concomitant evolutionary pressures for function and folding efficiency.

View Article: PubMed Central - PubMed

Affiliation: Astbury Centre for Structural Molecular Biology, University of Leeds, Mount Preston Street, Leeds LS2 9JT, UK.

ABSTRACT
Many proteins reach their native state through pathways involving the presence of folding intermediates. It is not clear whether this type of folding landscape results from insufficient evolutionary pressure to optimize folding efficiency, or arises from a conflict between functional and folding constraints. Here, using protein-engineering, ultra-rapid mixing and stopped-flow experiments combined with restrained molecular dynamics simulations, we characterize the transition state for the formation of the intermediate populated during the folding of the bacterial immunity protein, Im7, and the subsequent molecular steps leading to the native state. The results provide a comprehensive view of the folding process of this small protein. An analysis of the contributions of native and non-native interactions at different stages of folding reveals how the complexity of the folding landscape arises from concomitant evolutionary pressures for function and folding efficiency.

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Interaction patterns of selected residues in TS1, I, TS2 and N. (a) Number of atomic contacts (native and non-native) formed between residues Phe41, Tyr55, Trp75 and the other residues of Im7 in TS1, I, TS2, and N. (b) Structures illustrating the position of residues Phe41 (green), Tyr55 (magenta), and Trp75 (orange) in each ensemble.
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Figure 8: Interaction patterns of selected residues in TS1, I, TS2 and N. (a) Number of atomic contacts (native and non-native) formed between residues Phe41, Tyr55, Trp75 and the other residues of Im7 in TS1, I, TS2, and N. (b) Structures illustrating the position of residues Phe41 (green), Tyr55 (magenta), and Trp75 (orange) in each ensemble.

Mentions: The number of side chain-side chain interactions between Phe41, Tyr55 and Trp75 and all other residues in TS1, I, TS2 and N are shown in Fig. 7a. These profiles reveal that Trp75 makes a large number of non-native interactions with residues in regions 37-45 (Helix II) and 51-56 (Helix III) in the intermediate. Moreover, inspection of representative structures from each ensemble (Fig. 7b) suggests that the non-native interactions formed between Trp75 and side chains of residues in helix II hinder residues in helix III (represented here by Tyr55) from adopting their native position in which these residues dock against buried side chains of residues in helices II and IV. To investigate this mechanism further, the distribution of distances between Phe41 (helix II) and either I54 (helix III) or Trp75 (helix IV) was determined for I, TS2 and N (Supplementary Fig. S4c online). Whilst Phe41 is close to Ile54 in TS2 and N, this is not the case for the intermediate. In fact, in many conformations of the intermediate ensemble Trp75 is closer to Phe41 than is Ile54. These results confirm that residues in the C-terminal region of helix II form substantial non-native interactions with Trp75 in the intermediate, thereby inhibiting helix III from finding its native interaction partners and temporarily trapping Im7 in the intermediate state.


The mechanism of folding of Im7 reveals competition between functional and kinetic evolutionary constraints.

Friel CT, Smith DA, Vendruscolo M, Gsponer J, Radford SE - Nat. Struct. Mol. Biol. (2009)

Interaction patterns of selected residues in TS1, I, TS2 and N. (a) Number of atomic contacts (native and non-native) formed between residues Phe41, Tyr55, Trp75 and the other residues of Im7 in TS1, I, TS2, and N. (b) Structures illustrating the position of residues Phe41 (green), Tyr55 (magenta), and Trp75 (orange) in each ensemble.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2651959&req=5

Figure 8: Interaction patterns of selected residues in TS1, I, TS2 and N. (a) Number of atomic contacts (native and non-native) formed between residues Phe41, Tyr55, Trp75 and the other residues of Im7 in TS1, I, TS2, and N. (b) Structures illustrating the position of residues Phe41 (green), Tyr55 (magenta), and Trp75 (orange) in each ensemble.
Mentions: The number of side chain-side chain interactions between Phe41, Tyr55 and Trp75 and all other residues in TS1, I, TS2 and N are shown in Fig. 7a. These profiles reveal that Trp75 makes a large number of non-native interactions with residues in regions 37-45 (Helix II) and 51-56 (Helix III) in the intermediate. Moreover, inspection of representative structures from each ensemble (Fig. 7b) suggests that the non-native interactions formed between Trp75 and side chains of residues in helix II hinder residues in helix III (represented here by Tyr55) from adopting their native position in which these residues dock against buried side chains of residues in helices II and IV. To investigate this mechanism further, the distribution of distances between Phe41 (helix II) and either I54 (helix III) or Trp75 (helix IV) was determined for I, TS2 and N (Supplementary Fig. S4c online). Whilst Phe41 is close to Ile54 in TS2 and N, this is not the case for the intermediate. In fact, in many conformations of the intermediate ensemble Trp75 is closer to Phe41 than is Ile54. These results confirm that residues in the C-terminal region of helix II form substantial non-native interactions with Trp75 in the intermediate, thereby inhibiting helix III from finding its native interaction partners and temporarily trapping Im7 in the intermediate state.

Bottom Line: It is not clear whether this type of folding landscape results from insufficient evolutionary pressure to optimize folding efficiency, or arises from a conflict between functional and folding constraints.The results provide a comprehensive view of the folding process of this small protein.An analysis of the contributions of native and non-native interactions at different stages of folding reveals how the complexity of the folding landscape arises from concomitant evolutionary pressures for function and folding efficiency.

View Article: PubMed Central - PubMed

Affiliation: Astbury Centre for Structural Molecular Biology, University of Leeds, Mount Preston Street, Leeds LS2 9JT, UK.

ABSTRACT
Many proteins reach their native state through pathways involving the presence of folding intermediates. It is not clear whether this type of folding landscape results from insufficient evolutionary pressure to optimize folding efficiency, or arises from a conflict between functional and folding constraints. Here, using protein-engineering, ultra-rapid mixing and stopped-flow experiments combined with restrained molecular dynamics simulations, we characterize the transition state for the formation of the intermediate populated during the folding of the bacterial immunity protein, Im7, and the subsequent molecular steps leading to the native state. The results provide a comprehensive view of the folding process of this small protein. An analysis of the contributions of native and non-native interactions at different stages of folding reveals how the complexity of the folding landscape arises from concomitant evolutionary pressures for function and folding efficiency.

Show MeSH