Cell-wide analysis of protein thermal unfolding reveals determinants of thermostability

Abstract

Temperature-induced cell death is thought to be due to protein denaturation, but the determinants of thermal sensitivity of proteomes remain largely uncharacterized. We developed a structural proteomic strategy to measure protein thermostability on a proteome-wide scale and with domain-level resolution. We applied it to ,, , and human cells, yielding thermostability data for more than 8000 proteins. Our results (i) indicate that temperature-induced cellular collapse is due to the loss of a subset of proteins with key functions, (ii) shed light on the evolutionary conservation of protein and domain stability, and (iii) suggest that natively disordered proteins in a cell are less prevalent than predicted and (iv) that highly expressed proteins are stable because they are designed to tolerate translational errors that would lead to the accumulation of toxic misfolded species.

Ancestral diterpene cyclases show increased thermostability and substrate acceptance

“…As a result, there might be a bias to automatically select the most stabilizing residue for each position within a certain clade of the tree . High protein stability has recently been connected to higher abundance of soluble protein in the cell . This is in accordance with our findings, namely that the two oldest ancestral terpene cyclases are readily expressed in E. coli with up to four times higher protein yield compared to their modern counterpart (Fig.…”

“…The consensus enzyme was found to generate a lower yield of soluble protein of less than 1 mg·L −1 . These findings suggested that SvS‐A1 and SvS‐A2 are generally more stable proteins . To confirm this, we established melting curves for all enzymes by monitoring intrinsic tryptophan fluorescence at 330 and 350 nm using nanoDSF (Table ).…”

DnaJ chaperones contribute to canalization

“…Therefore, it is essential to investigate how fluctuations in temperature affect such proteins and how ER‐associated DnaJ chaperones rescue such disordered proteins under thermal stress. Recent advances in mass spectrometry analysis has enabled monitoring of protein structural change under various temperatures (Leuenberger et al, ). Applying such technology to reveal the interactome of these DnaJ chaperones will further test the molecular mechanisms underpinning canalization.…”

Emerging roles of allosteric modulators in the regulation of protein‐protein interactions (PPIs): A new paradigm for PPI drug discovery

“…There are various optimized versions of MS‐based methods, such as tandem affinity purification MS, affinity enrichment MS, and immunopurification MS, which supply comprehensive insights into the dynamic PPI networks . As a powerful tool for PPI detection, especially at the level of protein complexes, MS has been widely applied for research regarding the interactome and proteome . Despite these successes, there remain limitations for MS‐based approaches, such as the relatively large requirement for sample input, and further miniaturizations and optimizations are still underway .…”