(183h) Atomistic Simulations Reveal Phosphorylation-Dependent Conformational Shifts in the Disordered N-Terminus of c-Myc1-88

The transcription factor c-Myc is one of the most commonly dysregulated proteins in cancer, playing key roles in proliferation, apoptosis, and metabolism. Yet, despite decades of effort, c-Myc remains an undruggable target due to its intrinsically disordered nature and lack of conventional binding pockets. An alternative strategy involves promoting c-Myc degradation since its sustained overexpression drives oncogenic transcriptional programs and cellular toxicity. This process is regulated by hierarchical phosphorylation at serine 62 (S62) and threonine 58 (T58), followed by ubiquitin-dependent proteasomal degradation. The role of S62 phosphorylation has long been perplexing: while it was traditionally viewed as a proteostabilizing modification, it is also necessary to prime T58 phosphorylation by GSK3, both of which is essential for degradation. Recent findings have deepened this paradox by showing that S62 phosphorylation not only primes T58 but also directly increases the affinity of the T58 degron for Fbw7 and promotes Fbw7-mediated ubiquitylation and degradation of Myc. This dual role of S62—in both stabilizing and promoting degradation—highlights a more complex regulatory mechanism than previously appreciated, complicating efforts to target this pathway for therapeutic benefit .

At the more fundamental level, recent studies have shown that the structural effects of phosphorylation within intrinsically disordered proteins (IDPs) are both residue- and context-specific, making it challenging to generalize their regulatory impact. Specifically, phosphorylation can induce disorder-to-order transitions or modulate interaction hotspots—effects that are only beginning to be explored in atomistic detail.

In this work, we bridge these lines of inquiry by performing enhanced sampling molecular dynamics simulations of c-Myc^1-88, encompassing MB0, MB1, and the critical phosphodegron regions. This region includes S62 and T58 and is central to c-Myc degradation. By modeling the non-phosphorylated states, the singly phosphorylated S62 states, the singly phosphorylated T58 states, and the doubly phosphorylated S62/T58 states, we examine how conformational ensembles shift as a function of phosphorylation. Our simulations capture key non-phosphorylated states that overlap with NMR-characterized structural motifs, such as transient α-helices in MB1 (residues 27–38).

Strikingly, our simulations revealed measurable differences in compactness, secondary structure content, residue-level fluctuations, and intramolecular contact patterns among the unphosphorylated, and phosphorylated states of c-Myc^1-88. These distinctions suggest that phosphorylation induces specific conformational changes in this intrinsically disordered region, potentially influencing its recognition by degradation machinery.

By uncovering how hierarchical phosphorylation alters the conformational landscape of the N-terminal region of c-Myc, this work opens new directions for drug discovery targeting the degradation pathway of c-Myc. Our atomistic insights may aid in the rational design of molecules that modulate these phosphorylation-dependent states to enhance c-Myc turnover in cancer cells.