Conservation of function without conservation of amino acid sequence in intrinsically disordered transcriptional activation domains

Protein function is canonically believed to be more conserved than amino acid sequence, but this idea is only well supported in folded domains, where highly diverged sequences can fold into equivalent 3D structures. In contrast, intrinsically disordered protein regions (IDRs) do not fold into a stable 3D structure, thus it remains unknown when and how function is conserved for IDRs that experience rapid amino acid sequence divergence. As a model system for studying the evolution of IDRs, we examined transcriptional activation domains, the regions of transcription factors that bind to coactivator complexes. We systematically identified activation domains on 502 orthologs of the transcriptional activator Gcn4 spanning 600 MY of fungal evolution. We find that the central activation domain shows strong conservation of function without conservation of sequence. This conservation of function without conservation of sequence is facilitated by evolutionary turnover (gain and loss) of key acidic and aromatic residues, the positions most important for function. This high sequence flexibility of functional orthologs mirrors the physical flexibility of the activation domain coactivator interaction interface, suggesting that physical flexibility enables evolutionary plasticity. We propose that turnover of short functional elements, sometimes individual amino acids, is a general mechanism for conservation of function without conservation of sequence during IDR evolution.