Cellular environments modulate protein energy landscapes to drive important biology, where small perturbations are consequential for biological signaling, allostery, and other vital processes. The energetic effects of ubiquitination are interesting due to its potential influence on degradation by the 26S proteasome, which requires intrinsically flexible or unstructured initiation regions that many known proteasome substrates lack. We generated proteins with natively attached, isopeptide-linked ubiquitin in structured domains to assess the energetic changes contributed by ubiquitin and how such changes manifest at the proteasome. Ubiquitination at sensitive sites destabilizes the native structure, and thereby increases the rate of degradation for substrates containing unstructured initiation regions. Importantly, this ubiquitination can even induce those requisite regions in well-folded proteins for proteasomal engagement. Our results indicate a biophysical role of site-specific ubiquitination as a potential regulatory mechanism for energy-dependent substrate degradation.

Ubiquitination modulates a protein energy landscape site-specifically with consequences for proteasomal degradation