A new study suggests that misfolded proteins with near-native structures may escape the cell’s degradation machinery, revealing a possible blind spot in protein quality control.
Researchers led by Ed O’Brien at Penn State tested whether misfolding linked to non-covalent lasso entanglements (NCLEs) influences protein degradation. These knot-like structural features can misfold when a native entanglement fails to form, or when an extra non-native entanglement forms instead.
To connect structure with protein fate, the team integrated AlphaFold models with ubiquitin mass spectrometry (Ubq-MS) data from human fibroblasts, using lysine di-glycine peptide signatures to identify proteins tagged for degradation by the ubiquitin-proteasome system.
“Like a tiny factory, cells make proteins,” said O’Brien in a recent press release. “And like a factory, cells have quality control mechanisms to catch any errors on the production line. We’ve recently identified a new class of protein misfolding, and we were interested in if it had any impact on how the cellular quality control system maintains a balance of protein production, repair and recycling – protein homeostasis.”
Among proteins with high-quality AlphaFold structures, those containing native NCLEs were more likely to be tagged soon after synthesis: 38 percent were young and ubiquitinated, compared with 20 percent of non-entangled proteins. After controlling for protein length, native NCLEs were associated with a 93 percent increase in the odds of young-age ubiquitination.
The same pattern appeared across additional ubiquitination datasets, suggesting that the effect was not specific to a single proteomics experiment. In coarse-grained molecular dynamics simulations, entangled, young-ubiquitinated proteins were more than four times as likely to misfold with entanglement changes as size-matched, non-entangled proteins that were not ubiquitinated.
Not all misfolded entangled proteins, however, appeared to be targeted for degradation. Some that were not ubiquitinated showed a similar tendency to misfold, but adopted more native-like conformations and exposed less hydrophobic surface, which may make them harder for quality-control machinery to recognize.
“Sometimes a misfolded entanglement can be hidden deep within the structure of a protein, so that it isn’t visible to the quality control system,” said first author Yang Jiang. “These proteins may therefore evade degradation and persist in the cell despite being non-functional.”
Combining the proteomics and simulation results, the authors estimate that nearly half of misfolded entangled proteins may evade ubiquitin-proteasome degradation and persist in soluble, near-native, but less functional states. If these near-native misfolded proteins persist in cells, the authors suggest they could gradually disrupt protein homeostasis, with possible relevance to aging and disease.
“Protein misfolding is known to contribute to diseases like Alzheimer’s and Huntington’s,” O’Brien said. “Therefore, increasing our understanding of the basic biology underlying this novel class of misfolding could lead to the identification of new disease origins and treatments.”
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