Proteostasis describes the management of proteome foldedness and involves an extensive quality control network that oversees the life of every protein from synthesis, to its folding, transport and ultimately degradation.  Proteostasis becomes unbalanced during neurodegenerative disease, which correlates with the abnormal deposition of protein aggregates that are known byproducts of protein misfolding.  Our laboratory is developing new strategies to probe how proteome foldedness and aggregation state changes upon proteostasis stress and determine which proteins are selectively vulnerable to misfolding and aggregation. One approach involves using a fluorigenic chemical probe, tetraphenylethene-maleimide (TPE-MI) to monitor cysteine reactivity.  Free cys residues are typically buried in the core of globular proteins and we hypothesized that an accumulation of unfolded proteins would expose more cys-thiols to the solvent and enable greater reactivity to TPE-MI.  TPE-MI can indeed capture a snapshot of the balance of unfolded protein relative to folded states in intact live cells (Chen et al. 2017, Nat. Comm.). Here we describe our approach and its validation with purified proteins and is application in several experimental models of cell stress.  We will also describe our efforts to quantitate the changes in proteome cys reactivity upon a panel of proteostasis stresses using mass spectrometry and in mapping which proteins are selectively altered in cys reactivity.    <br/>

Probing thiol-reactivity to monitor proteome foldedness and conformational change under proteostasis stress

2019 Fall National Meeting

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