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Transient structure and dynamics in the disordered c-Myc transactivation domain affect Bin1 binding.

Andresen C, Helander S, Lemak A, Farès C, Csizmok V, Carlsson J, Penn LZ, Forman-Kay JD, Arrowsmith CH, Lundström P, Sunnerhagen M - Nucleic Acids Res. (2012)

Bottom Line: We have characterized in detail the intrinsically disordered properties of Myc-1-88, where hierarchical phosphorylation of S62 and T58 regulates activation and destruction of the Myc protein.Binding of Bin1 to Myc-1-88 as assayed by NMR and surface plasmon resonance (SPR) revealed primary binding to the S62 region in a dynamically disordered and multivalent complex, accompanied by population shifts leading to altered intramolecular conformational dynamics.These findings expand the increasingly recognized concept of intrinsically disordered regions mediating transient interactions to Myc, a key transcriptional regulator of major medical importance, and have important implications for further understanding its multifaceted role in gene regulation.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Biotechnology, Department of Physics, Chemistry and Biology, Linköping University, SE-58183 Linköping, Sweden.

ABSTRACT
The crucial role of Myc as an oncoprotein and as a key regulator of cell growth makes it essential to understand the molecular basis of Myc function. The N-terminal region of c-Myc coordinates a wealth of protein interactions involved in transformation, differentiation and apoptosis. We have characterized in detail the intrinsically disordered properties of Myc-1-88, where hierarchical phosphorylation of S62 and T58 regulates activation and destruction of the Myc protein. By nuclear magnetic resonance (NMR) chemical shift analysis, relaxation measurements and NOE analysis, we show that although Myc occupies a very heterogeneous conformational space, we find transiently structured regions in residues 22-33 and in the Myc homology box I (MBI; residues 45-65); both these regions are conserved in other members of the Myc family. Binding of Bin1 to Myc-1-88 as assayed by NMR and surface plasmon resonance (SPR) revealed primary binding to the S62 region in a dynamically disordered and multivalent complex, accompanied by population shifts leading to altered intramolecular conformational dynamics. These findings expand the increasingly recognized concept of intrinsically disordered regions mediating transient interactions to Myc, a key transcriptional regulator of major medical importance, and have important implications for further understanding its multifaceted role in gene regulation.

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15N Relaxation parameters of free Myc-1–88 (open circles) and Myc-1–88 bound to Bin1–SH3 in a 1:1.5 ratio (filled circles). (A) {1H}-15N-NOE. (B) R1 relaxation rate constants. (C) R2 relaxation rate constants.
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gks263-F7: 15N Relaxation parameters of free Myc-1–88 (open circles) and Myc-1–88 bound to Bin1–SH3 in a 1:1.5 ratio (filled circles). (A) {1H}-15N-NOE. (B) R1 relaxation rate constants. (C) R2 relaxation rate constants.

Mentions: To investigate whether Bin1–SH3 binding results in reduced intrinsic disorder within the Myc binding site(s), the dynamic properties of Myc-1–88 on a rapid time scale (ps–ns) were assayed in the absence and presence of ligand binding. Backbone 15N relaxation experiments (R1, R1ρ, NOE) were performed on the apo and complex forms of Myc-1–88 (Figure 7). Measurements were made at 1.5 equivalents of Bin1–SH3 relative to Myc in order to achieve high saturation of the primary site while limiting binding to second, lower affinity site(s). Peaks with overlap leading to difficulties in fitting the data were not included in the evaluation.Figure 7.


Transient structure and dynamics in the disordered c-Myc transactivation domain affect Bin1 binding.

Andresen C, Helander S, Lemak A, Farès C, Csizmok V, Carlsson J, Penn LZ, Forman-Kay JD, Arrowsmith CH, Lundström P, Sunnerhagen M - Nucleic Acids Res. (2012)

15N Relaxation parameters of free Myc-1–88 (open circles) and Myc-1–88 bound to Bin1–SH3 in a 1:1.5 ratio (filled circles). (A) {1H}-15N-NOE. (B) R1 relaxation rate constants. (C) R2 relaxation rate constants.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3401448&req=5

gks263-F7: 15N Relaxation parameters of free Myc-1–88 (open circles) and Myc-1–88 bound to Bin1–SH3 in a 1:1.5 ratio (filled circles). (A) {1H}-15N-NOE. (B) R1 relaxation rate constants. (C) R2 relaxation rate constants.
Mentions: To investigate whether Bin1–SH3 binding results in reduced intrinsic disorder within the Myc binding site(s), the dynamic properties of Myc-1–88 on a rapid time scale (ps–ns) were assayed in the absence and presence of ligand binding. Backbone 15N relaxation experiments (R1, R1ρ, NOE) were performed on the apo and complex forms of Myc-1–88 (Figure 7). Measurements were made at 1.5 equivalents of Bin1–SH3 relative to Myc in order to achieve high saturation of the primary site while limiting binding to second, lower affinity site(s). Peaks with overlap leading to difficulties in fitting the data were not included in the evaluation.Figure 7.

Bottom Line: We have characterized in detail the intrinsically disordered properties of Myc-1-88, where hierarchical phosphorylation of S62 and T58 regulates activation and destruction of the Myc protein.Binding of Bin1 to Myc-1-88 as assayed by NMR and surface plasmon resonance (SPR) revealed primary binding to the S62 region in a dynamically disordered and multivalent complex, accompanied by population shifts leading to altered intramolecular conformational dynamics.These findings expand the increasingly recognized concept of intrinsically disordered regions mediating transient interactions to Myc, a key transcriptional regulator of major medical importance, and have important implications for further understanding its multifaceted role in gene regulation.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Biotechnology, Department of Physics, Chemistry and Biology, Linköping University, SE-58183 Linköping, Sweden.

ABSTRACT
The crucial role of Myc as an oncoprotein and as a key regulator of cell growth makes it essential to understand the molecular basis of Myc function. The N-terminal region of c-Myc coordinates a wealth of protein interactions involved in transformation, differentiation and apoptosis. We have characterized in detail the intrinsically disordered properties of Myc-1-88, where hierarchical phosphorylation of S62 and T58 regulates activation and destruction of the Myc protein. By nuclear magnetic resonance (NMR) chemical shift analysis, relaxation measurements and NOE analysis, we show that although Myc occupies a very heterogeneous conformational space, we find transiently structured regions in residues 22-33 and in the Myc homology box I (MBI; residues 45-65); both these regions are conserved in other members of the Myc family. Binding of Bin1 to Myc-1-88 as assayed by NMR and surface plasmon resonance (SPR) revealed primary binding to the S62 region in a dynamically disordered and multivalent complex, accompanied by population shifts leading to altered intramolecular conformational dynamics. These findings expand the increasingly recognized concept of intrinsically disordered regions mediating transient interactions to Myc, a key transcriptional regulator of major medical importance, and have important implications for further understanding its multifaceted role in gene regulation.

Show MeSH
Related in: MedlinePlus