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Structure, function, and dynamics of the dimerization and DNA-binding domain of oncogenic transcription factor v-Myc.
J Mol Biol. 2001 Apr 13; 307(5):1395-410.JM

Abstract

The protein product (c-Myc) of the protooncogene c-myc is a transcriptional regulator playing a key role in cellular growth, differentiation, and apoptosis. Deregulated myc genes, like the transduced retroviral v-myc allele, are oncogenic and cause cell transformation. The C-terminal bHLHZip domain of v-Myc, encompassing protein dimerization (helix-loop-helix, leucine zipper) and DNA contact (basic region) surfaces, was expressed in bacteria as a highly soluble p15(v-myc)recombinant protein. Dissociation constants (K(d)) for the heterodimer formed with the recombinant bHLHZip domain of the Myc binding partner Max (p14(max)) and for the Myc-Max-DNA complex were estimated using circular dichroism (CD) spectroscopy and quantitative electrophoretic mobility shift assay (EMSA). Multi-dimensional NMR spectroscopy was used to characterize the solution structural and dynamic properties of the v-Myc bHLHZip domain. Significant secondary chemical shifts indicate the presence of two separated alpha-helical regions. The C-terminal leucine zipper region forms a compact alpha-helix, while the N-terminal basic region exhibits conformational averaging with substantial alpha-helical content. Both helices lack stabilizing tertiary side-chain interactions and represent exceptional examples for loosely coupled secondary structural segments in a native protein. These results and CD thermal denaturation data indicate a monomeric state of the v-Myc bHLHZip domain. The (15)N relaxation data revealed backbone mobilities which corroborate the existence of a partially folded state, and suggest a "beads-on-a-string" motional behaviour of the v-Myc bHLHZip domain in solution. The preformation of alpha-helical regions was confirmed by CD thermal denaturation studies, and quantification of the entropy changes caused by the hydrophobic effect and the reduction of conformational entropy upon protein dimerization. The restricted conformational space of the v-Myc bHLHZip domain reduces the entropy penalty associated with heterodimerization and allows rapid and accurate recognition by the authentic Myc binding partner Max.

Authors+Show Affiliations

Institute of Organic Chemistry andNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

11292350

Citation

Fieber, W, et al. "Structure, Function, and Dynamics of the Dimerization and DNA-binding Domain of Oncogenic Transcription Factor V-Myc." Journal of Molecular Biology, vol. 307, no. 5, 2001, pp. 1395-410.
Fieber W, Schneider ML, Matt T, et al. Structure, function, and dynamics of the dimerization and DNA-binding domain of oncogenic transcription factor v-Myc. J Mol Biol. 2001;307(5):1395-410.
Fieber, W., Schneider, M. L., Matt, T., Kräutler, B., Konrat, R., & Bister, K. (2001). Structure, function, and dynamics of the dimerization and DNA-binding domain of oncogenic transcription factor v-Myc. Journal of Molecular Biology, 307(5), 1395-410.
Fieber W, et al. Structure, Function, and Dynamics of the Dimerization and DNA-binding Domain of Oncogenic Transcription Factor V-Myc. J Mol Biol. 2001 Apr 13;307(5):1395-410. PubMed PMID: 11292350.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Structure, function, and dynamics of the dimerization and DNA-binding domain of oncogenic transcription factor v-Myc. AU - Fieber,W, AU - Schneider,M L, AU - Matt,T, AU - Kräutler,B, AU - Konrat,R, AU - Bister,K, PY - 2001/4/9/pubmed PY - 2001/5/22/medline PY - 2001/4/9/entrez SP - 1395 EP - 410 JF - Journal of molecular biology JO - J Mol Biol VL - 307 IS - 5 N2 - The protein product (c-Myc) of the protooncogene c-myc is a transcriptional regulator playing a key role in cellular growth, differentiation, and apoptosis. Deregulated myc genes, like the transduced retroviral v-myc allele, are oncogenic and cause cell transformation. The C-terminal bHLHZip domain of v-Myc, encompassing protein dimerization (helix-loop-helix, leucine zipper) and DNA contact (basic region) surfaces, was expressed in bacteria as a highly soluble p15(v-myc)recombinant protein. Dissociation constants (K(d)) for the heterodimer formed with the recombinant bHLHZip domain of the Myc binding partner Max (p14(max)) and for the Myc-Max-DNA complex were estimated using circular dichroism (CD) spectroscopy and quantitative electrophoretic mobility shift assay (EMSA). Multi-dimensional NMR spectroscopy was used to characterize the solution structural and dynamic properties of the v-Myc bHLHZip domain. Significant secondary chemical shifts indicate the presence of two separated alpha-helical regions. The C-terminal leucine zipper region forms a compact alpha-helix, while the N-terminal basic region exhibits conformational averaging with substantial alpha-helical content. Both helices lack stabilizing tertiary side-chain interactions and represent exceptional examples for loosely coupled secondary structural segments in a native protein. These results and CD thermal denaturation data indicate a monomeric state of the v-Myc bHLHZip domain. The (15)N relaxation data revealed backbone mobilities which corroborate the existence of a partially folded state, and suggest a "beads-on-a-string" motional behaviour of the v-Myc bHLHZip domain in solution. The preformation of alpha-helical regions was confirmed by CD thermal denaturation studies, and quantification of the entropy changes caused by the hydrophobic effect and the reduction of conformational entropy upon protein dimerization. The restricted conformational space of the v-Myc bHLHZip domain reduces the entropy penalty associated with heterodimerization and allows rapid and accurate recognition by the authentic Myc binding partner Max. SN - 0022-2836 UR - https://www.unboundmedicine.com/medline/citation/11292350/Structure_function_and_dynamics_of_the_dimerization_and_DNA_binding_domain_of_oncogenic_transcription_factor_v_Myc_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0022-2836(01)94537-1 DB - PRIME DP - Unbound Medicine ER -