Thermodynamics of the high-affinity interaction of TCF4 with beta-catenin.
The formation of a complex between beta-catenin and members of the TCF/LEF family of high-mobility group proteins is a key regulatory event in the wnt-signaling pathway, essential for embryonal development as well as the growth of normal and malignant colon epithelium. We have characterized the bind...
| Main Authors: | , , , , , , , , , , |
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| Format: | Journal article |
| Language: | English |
| Published: |
2001
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| _version_ | 1797084489989488640 |
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| author | Knapp, S Zamai, M Volpi, D Nardese, V Avanzi, N Breton, J Plyte, S Flocco, M Marconi, M Isacchi, A Caiolfa, V |
| author_facet | Knapp, S Zamai, M Volpi, D Nardese, V Avanzi, N Breton, J Plyte, S Flocco, M Marconi, M Isacchi, A Caiolfa, V |
| author_sort | Knapp, S |
| collection | OXFORD |
| description | The formation of a complex between beta-catenin and members of the TCF/LEF family of high-mobility group proteins is a key regulatory event in the wnt-signaling pathway, essential for embryonal development as well as the growth of normal and malignant colon epithelium. We have characterized the binding of TCF4 to human beta-catenin by steady-state intrinsic fluorescence quenching experiments, surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC). Binding studies in solution and in heterogeneous phase showed that TCF4 binds reversibly to beta-catenin with an affinity (KB) of 3(+/-1) 10(8) M(-1). Site-directed mutagenesis, together with calorimetric measurements, revealed that residue D16 in TCF4 plays a crucial role in high-affinity binding. Mutation of this residue to alanine resulted in a decrease of KB by two orders of magnitude as well as a significant reduction in binding enthalpy. Binding of TCF4 to beta-catenin gave rise to a large negative enthalpy change at 25 degrees C (-29.7 kcal/mol). Binding enthalpies were strongly temperature dependent, which resulted in the determination of a large heat capacity change upon binding of -1.5 kcal/(mol K). The molecular events that take place upon complex formation are discussed using the measured thermodynamic data together with the crystal structure of the beta-catenin arm repeat region/TCF complex. |
| first_indexed | 2024-03-07T01:55:50Z |
| format | Journal article |
| id | oxford-uuid:9bae0f90-f888-4d24-9abf-53d03b0312d6 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-07T01:55:50Z |
| publishDate | 2001 |
| record_format | dspace |
| spelling | oxford-uuid:9bae0f90-f888-4d24-9abf-53d03b0312d62022-03-27T00:30:28ZThermodynamics of the high-affinity interaction of TCF4 with beta-catenin.Journal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:9bae0f90-f888-4d24-9abf-53d03b0312d6EnglishSymplectic Elements at Oxford2001Knapp, SZamai, MVolpi, DNardese, VAvanzi, NBreton, JPlyte, SFlocco, MMarconi, MIsacchi, ACaiolfa, VThe formation of a complex between beta-catenin and members of the TCF/LEF family of high-mobility group proteins is a key regulatory event in the wnt-signaling pathway, essential for embryonal development as well as the growth of normal and malignant colon epithelium. We have characterized the binding of TCF4 to human beta-catenin by steady-state intrinsic fluorescence quenching experiments, surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC). Binding studies in solution and in heterogeneous phase showed that TCF4 binds reversibly to beta-catenin with an affinity (KB) of 3(+/-1) 10(8) M(-1). Site-directed mutagenesis, together with calorimetric measurements, revealed that residue D16 in TCF4 plays a crucial role in high-affinity binding. Mutation of this residue to alanine resulted in a decrease of KB by two orders of magnitude as well as a significant reduction in binding enthalpy. Binding of TCF4 to beta-catenin gave rise to a large negative enthalpy change at 25 degrees C (-29.7 kcal/mol). Binding enthalpies were strongly temperature dependent, which resulted in the determination of a large heat capacity change upon binding of -1.5 kcal/(mol K). The molecular events that take place upon complex formation are discussed using the measured thermodynamic data together with the crystal structure of the beta-catenin arm repeat region/TCF complex. |
| spellingShingle | Knapp, S Zamai, M Volpi, D Nardese, V Avanzi, N Breton, J Plyte, S Flocco, M Marconi, M Isacchi, A Caiolfa, V Thermodynamics of the high-affinity interaction of TCF4 with beta-catenin. |
| title | Thermodynamics of the high-affinity interaction of TCF4 with beta-catenin. |
| title_full | Thermodynamics of the high-affinity interaction of TCF4 with beta-catenin. |
| title_fullStr | Thermodynamics of the high-affinity interaction of TCF4 with beta-catenin. |
| title_full_unstemmed | Thermodynamics of the high-affinity interaction of TCF4 with beta-catenin. |
| title_short | Thermodynamics of the high-affinity interaction of TCF4 with beta-catenin. |
| title_sort | thermodynamics of the high affinity interaction of tcf4 with beta catenin |
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