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Binding-induced folding of a natively unstructured transcription factor.

Turjanski AG, Gutkind JS, Best RB, Hummer G - PLoS Comput. Biol. (2008)

Bottom Line: Interestingly, increasing the amount of structure in the unbound pKID reduces the rate of binding, suggesting a "fly-casting"-like process.We find that the inclusion of attractive non-native interactions results in the formation of non-specific encounter complexes that enhance the on-rate of binding, but do not significantly change the binding mechanism.The simulations are in general agreement with the results of a recently reported nuclear magnetic resonance study, and aid in the interpretation of the experimental binding kinetics.

View Article: PubMed Central - PubMed

Affiliation: Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, Maryland, United States of America.

ABSTRACT
Transcription factors are central components of the intracellular regulatory networks that control gene expression. An increasingly recognized phenomenon among human transcription factors is the formation of structure upon target binding. Here, we study the folding and binding of the pKID domain of CREB to the KIX domain of the co-activator CBP. Our simulations of a topology-based Gō-type model predict a coupled folding and binding mechanism, and the existence of partially bound intermediates. From transition-path and Phi-value analyses, we find that the binding transition state resembles the unstructured state in solution, implying that CREB becomes structured only after committing to binding. A change of structure following binding is reminiscent of an induced-fit mechanism and contrasts with models in which binding occurs to pre-structured conformations that exist in the unbound state at equilibrium. Interestingly, increasing the amount of structure in the unbound pKID reduces the rate of binding, suggesting a "fly-casting"-like process. We find that the inclusion of attractive non-native interactions results in the formation of non-specific encounter complexes that enhance the on-rate of binding, but do not significantly change the binding mechanism. Our study helps explain how being unstructured can confer an advantage in protein target recognition. The simulations are in general agreement with the results of a recently reported nuclear magnetic resonance study, and aid in the interpretation of the experimental binding kinetics.

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Φ value analysis for the transition state.Φ values for the folding/binding transition state for the residues of pKID forming native contacts with KIX. Red squares indicate amino acids belonging to helix αA, green circles indicate amino acids belonging to helix αB, and black triangles show Φ values for residues without secondary structure.
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pcbi-1000060-g006: Φ value analysis for the transition state.Φ values for the folding/binding transition state for the residues of pKID forming native contacts with KIX. Red squares indicate amino acids belonging to helix αA, green circles indicate amino acids belonging to helix αB, and black triangles show Φ values for residues without secondary structure.

Mentions: In analogy to protein folding [34],[35] we further characterized the TS by calculating Φ values for each amino acid residue of pKID that is involved in native contacts with KIX. We estimated Φ values by using a simple definition in which only native contacts are counted, Φ = Qxc(TS)/Qxc(bound) [34],[36],[37], where Qxc is the number of native contacts in the respective state between protein 1 and residue x of protein 2. Φ values have been extensively used for determining key residues in the folding of proteins and remarkable agreement has been found between theoretical and experimental studies [34],[38],[39]. Φ-value analysis should quantify the amount of intermolecular structure formed at each site in the transition state. The calculated Φ values for the coupled folding-and-binding transition of pKID are shown in Figure 6. According to our analysis, only residues of helix αB seem to be important for TS formation, with the highest Φ values being found for Leu141 and the neighboring amino acids Asp140 and Ser142, implying that hydrophobic interactions involving amino acids at the center of the αB region govern the association mechanism. A critical role for Leu141, which is deeply bound into the hydrophobic groove of KIX, has been suggested based on the results of mutating Tyr650 of CBP which is located in the wall of the hydrophobic groove [11]. Mutation to Ala resulted in a ∼12-fold decrease in the binding constant [11]. Our study suggests that corresponding kinetic measurements on this mutant should be insightful.


Binding-induced folding of a natively unstructured transcription factor.

Turjanski AG, Gutkind JS, Best RB, Hummer G - PLoS Comput. Biol. (2008)

Φ value analysis for the transition state.Φ values for the folding/binding transition state for the residues of pKID forming native contacts with KIX. Red squares indicate amino acids belonging to helix αA, green circles indicate amino acids belonging to helix αB, and black triangles show Φ values for residues without secondary structure.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2289845&req=5

pcbi-1000060-g006: Φ value analysis for the transition state.Φ values for the folding/binding transition state for the residues of pKID forming native contacts with KIX. Red squares indicate amino acids belonging to helix αA, green circles indicate amino acids belonging to helix αB, and black triangles show Φ values for residues without secondary structure.
Mentions: In analogy to protein folding [34],[35] we further characterized the TS by calculating Φ values for each amino acid residue of pKID that is involved in native contacts with KIX. We estimated Φ values by using a simple definition in which only native contacts are counted, Φ = Qxc(TS)/Qxc(bound) [34],[36],[37], where Qxc is the number of native contacts in the respective state between protein 1 and residue x of protein 2. Φ values have been extensively used for determining key residues in the folding of proteins and remarkable agreement has been found between theoretical and experimental studies [34],[38],[39]. Φ-value analysis should quantify the amount of intermolecular structure formed at each site in the transition state. The calculated Φ values for the coupled folding-and-binding transition of pKID are shown in Figure 6. According to our analysis, only residues of helix αB seem to be important for TS formation, with the highest Φ values being found for Leu141 and the neighboring amino acids Asp140 and Ser142, implying that hydrophobic interactions involving amino acids at the center of the αB region govern the association mechanism. A critical role for Leu141, which is deeply bound into the hydrophobic groove of KIX, has been suggested based on the results of mutating Tyr650 of CBP which is located in the wall of the hydrophobic groove [11]. Mutation to Ala resulted in a ∼12-fold decrease in the binding constant [11]. Our study suggests that corresponding kinetic measurements on this mutant should be insightful.

Bottom Line: Interestingly, increasing the amount of structure in the unbound pKID reduces the rate of binding, suggesting a "fly-casting"-like process.We find that the inclusion of attractive non-native interactions results in the formation of non-specific encounter complexes that enhance the on-rate of binding, but do not significantly change the binding mechanism.The simulations are in general agreement with the results of a recently reported nuclear magnetic resonance study, and aid in the interpretation of the experimental binding kinetics.

View Article: PubMed Central - PubMed

Affiliation: Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, Maryland, United States of America.

ABSTRACT
Transcription factors are central components of the intracellular regulatory networks that control gene expression. An increasingly recognized phenomenon among human transcription factors is the formation of structure upon target binding. Here, we study the folding and binding of the pKID domain of CREB to the KIX domain of the co-activator CBP. Our simulations of a topology-based Gō-type model predict a coupled folding and binding mechanism, and the existence of partially bound intermediates. From transition-path and Phi-value analyses, we find that the binding transition state resembles the unstructured state in solution, implying that CREB becomes structured only after committing to binding. A change of structure following binding is reminiscent of an induced-fit mechanism and contrasts with models in which binding occurs to pre-structured conformations that exist in the unbound state at equilibrium. Interestingly, increasing the amount of structure in the unbound pKID reduces the rate of binding, suggesting a "fly-casting"-like process. We find that the inclusion of attractive non-native interactions results in the formation of non-specific encounter complexes that enhance the on-rate of binding, but do not significantly change the binding mechanism. Our study helps explain how being unstructured can confer an advantage in protein target recognition. The simulations are in general agreement with the results of a recently reported nuclear magnetic resonance study, and aid in the interpretation of the experimental binding kinetics.

Show MeSH