BPS2026 – A widespread protein misfolding mechanism is differentially rescued by chaperones based on gene essentiality

A new class of protein misfolding, involving changes in non-covalent lasso entanglement (NCLE) status, has been proposed based on simulations and biochemical assays. However, those studies were limited to a small number of proteins. Here, we detect hallmarks of these misfolded states in mixtures of hundreds of proteins by combining E. coli proteome-wide limited-proteolysis mass spectrometry (MS) data with structural data sets of protein native structures. We demonstrate that proteins containing native NCLEs are twice as likely to misfold, predominantly in regions where these NCLEs naturally occur. Surprisingly, the chaperones DnaK and GroEL do not typically correct this misfolding, except in the case of essential proteins. A statistical analysis reveals this differential rescue activity is related to weaker loop-closing contacts in the NCLEs of essential proteins, suggesting that misfolding involving these loops is easier to rectify by chaperones. Molecular simulations indicate a mechanism where premature NCLE loop closure, prior to proper placement of the threading segment, leads to persistent misfolded states. This mechanism explains why, in the mass spectrometry data, proteins containing native NCLEs are more likely to misfold and why misfolding occurs more frequently in native NCLE regions. These results suggest that NCLE misfolding is widespread, that such misfolded states in non-essential proteins can bypass the refolding action of chaperones, and that some protein sequences may have evolved to allow chaperone rescue from this class of misfolding.