N and C-terminal Sub-regions in the c-Myc Transactivation Region and their Joint Role in Creating Versatility in Folding and Binding

• Published in: Journal of molecular biology 2005-02, Vol.346 (1), p.175-189
• Main Authors: Fladvad, Malin, Zhou, Kaisong, Moshref, Ahmad, Pursglove, Sharon, Säfsten, Pär, Sunnerhagen, Maria
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; binding; Circular Dichroism; Cloning, Molecular; Humans; Medicin och hälsovetenskap; Molecular Sequence Data; molten globule; NATURAL SCIENCES; NATURVETENSKAP; Protein Binding; Protein Folding; Protein Structure, Secondary; Protein Structure, Tertiary; Proto-Oncogene Proteins c-myc - chemistry; Proto-Oncogene Proteins c-myc - genetics; Proto-Oncogene Proteins c-myc - metabolism; Sequence Alignment; Spectrometry, Fluorescence; surface plasmon resonance; transactivation domain; Transcriptional Activation
• ISSN: 0022-2836; 1089-8638; 1089-8638
• DOI: 10.1016/j.jmb.2004.11.029

The proto-oncogene c- myc governs the expression of a number of genes targeting cell growth and apoptosis, and its expression levels are distorted in many cancer forms. The current investigation presents an analysis by proteolysis, circular dichroism, fluorescence and Biacore of the folding and ligand-binding properties of the N-terminal transactivation domain (TAD) in the c-Myc protein. A c-Myc sub-region comprising residues 1–167 (Myc 1–167) has been investigated that includes the unstructured c-Myc transactivation domain (TAD, residues 1–143) together with a C-terminal segment, which appears to promote increased folding. Myc 1–167 is partly helical, binds both to the target proteins Myc modulator-1 (MM-1) and TATA box-binding protein (TBP), and displays the characteristics of a molten globule. Limited proteolysis divides Myc 1–167 in two halves, by cleaving in a predicted linker region between two hotspot mutation regions: Myc box I (MBI) and Myc box II (MBII). The N-terminal half (Myc 1–88) is unfolded and does not alone bind to target proteins, whereas the C-terminal half (Myc 92–167) has a partly helical fold and specifically binds both MM-1 and TBP. Although this might suggest a bipartite organization in the c-Myc TAD, none of the N and C-terminal fragments bind target protein with as high affinity as the entire Myc 1–167, or display molten globule properties. Furthermore, merely linking the MBI with the C-terminal region, in Myc 38–167, is not sufficient to achieve binding and folding properties as in Myc 1–167. Thus, the entire N and C-terminal regions of c-Myc TAD act in concert to achieve high specificity and affinity to two structurally and functionally orthogonal target proteins, TBP and MM-1, possibly through a mechanism involving molten globule formation. This hints towards understanding how binding of a range of targets can be accomplished to a single transactivation domain.