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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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Structural characteristics of the transition state ensemble.(A) and (B) show the probability density function (pdf) of intramolecular native contacts of helix αA (QαA) and helix αB (QαB) of pKID for the unbound state, TS, intermediates IA and IB, and bound states. (C) and (D) show the probability density functions of native contact fractions QCA and QCB between KIX and helices αA and αB of pKID, respectively, in the bound and transition states.
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pcbi-1000060-g005: Structural characteristics of the transition state ensemble.(A) and (B) show the probability density function (pdf) of intramolecular native contacts of helix αA (QαA) and helix αB (QαB) of pKID for the unbound state, TS, intermediates IA and IB, and bound states. (C) and (D) show the probability density functions of native contact fractions QCA and QCB between KIX and helices αA and αB of pKID, respectively, in the bound and transition states.

Mentions: A recently published study used NMR relaxation-dispersion experiments [23] to infer the formation of a single binding intermediate. Interestingly, the rates of interconversion of the intermediate and the bound state were fitted separately to the relaxation-dispersion curves for certain “clusters” of residues, with marked differences between the rate coefficients for clusters belonging to αA and those belonging to αB (see Table 1 in [23]). The rate of conversion from the intermediate state to the fully-bound state was more than four times faster for clusters of residues belonging to αB than for those belonging to αA, whereas the rate for conversion from the fully-bound to the intermediate state was faster for residues in αA than αB, an unexpected result for a strictly two-state system. Our simulations suggest a simple mechanism which explains this finding: the intermediate state actually consists of two sub-states, one (IA) with helix αB folded and bound, and helix αA unbound and only partially formed; the other (IB) with αA bound, and αB unbound. One can associate the rates from helix αA with the interconversion of intermediate IA with the bound state, and those from helix αB with the interconversion of IB and the bound state. From the ratio of these experimentally measured rates (Table 1 [23]), it follows that in steady state the population of IA is about 20 times larger than that of IB. Indeed, we find in the simulations that the population of IA is significantly larger than that of IB (Figure 4 and Figure S1), with a population ratio of ∼70, thus differing by a factor of about 3.5 from experiment (or about kBT ln 3.5∼0.75 kcal/mol in the free energy). Structurally, based on Figure 5A and 5B, we predict ∼80% intramolecular helical contacts for residues on áA, and ∼70% for áB. This is in reasonable agreement with the experimental estimates of ∼90% folded for áA and ∼70% folded for áB, inferred from NMR chemical shift differences [23]. (Note that the measured chemical shift differences of the different residues report on the structure in the respective intermediates: IA for residues in αA, and IB in αB, not the population-weighted average.) Overall, we find nearly quantitative agreement with experiment, with respect to both the equilibrium populations of the two intermediates and their structural characteristics. Thus our work is a true prediction of binding mechanism from simulation.


Binding-induced folding of a natively unstructured transcription factor.

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

Structural characteristics of the transition state ensemble.(A) and (B) show the probability density function (pdf) of intramolecular native contacts of helix αA (QαA) and helix αB (QαB) of pKID for the unbound state, TS, intermediates IA and IB, and bound states. (C) and (D) show the probability density functions of native contact fractions QCA and QCB between KIX and helices αA and αB of pKID, respectively, in the bound and transition states.
© Copyright Policy
Related In: Results  -  Collection

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

pcbi-1000060-g005: Structural characteristics of the transition state ensemble.(A) and (B) show the probability density function (pdf) of intramolecular native contacts of helix αA (QαA) and helix αB (QαB) of pKID for the unbound state, TS, intermediates IA and IB, and bound states. (C) and (D) show the probability density functions of native contact fractions QCA and QCB between KIX and helices αA and αB of pKID, respectively, in the bound and transition states.
Mentions: A recently published study used NMR relaxation-dispersion experiments [23] to infer the formation of a single binding intermediate. Interestingly, the rates of interconversion of the intermediate and the bound state were fitted separately to the relaxation-dispersion curves for certain “clusters” of residues, with marked differences between the rate coefficients for clusters belonging to αA and those belonging to αB (see Table 1 in [23]). The rate of conversion from the intermediate state to the fully-bound state was more than four times faster for clusters of residues belonging to αB than for those belonging to αA, whereas the rate for conversion from the fully-bound to the intermediate state was faster for residues in αA than αB, an unexpected result for a strictly two-state system. Our simulations suggest a simple mechanism which explains this finding: the intermediate state actually consists of two sub-states, one (IA) with helix αB folded and bound, and helix αA unbound and only partially formed; the other (IB) with αA bound, and αB unbound. One can associate the rates from helix αA with the interconversion of intermediate IA with the bound state, and those from helix αB with the interconversion of IB and the bound state. From the ratio of these experimentally measured rates (Table 1 [23]), it follows that in steady state the population of IA is about 20 times larger than that of IB. Indeed, we find in the simulations that the population of IA is significantly larger than that of IB (Figure 4 and Figure S1), with a population ratio of ∼70, thus differing by a factor of about 3.5 from experiment (or about kBT ln 3.5∼0.75 kcal/mol in the free energy). Structurally, based on Figure 5A and 5B, we predict ∼80% intramolecular helical contacts for residues on áA, and ∼70% for áB. This is in reasonable agreement with the experimental estimates of ∼90% folded for áA and ∼70% folded for áB, inferred from NMR chemical shift differences [23]. (Note that the measured chemical shift differences of the different residues report on the structure in the respective intermediates: IA for residues in αA, and IB in αB, not the population-weighted average.) Overall, we find nearly quantitative agreement with experiment, with respect to both the equilibrium populations of the two intermediates and their structural characteristics. Thus our work is a true prediction of binding mechanism from simulation.

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