Abstract
The Hsp70 chaperone system is integrated into a myriad of biochemical processes that are critical for cellular proteostasis. Although detailed pictures of Hsp70 bound with peptides have emerged, correspondingly detailed structural information on complexes with folding-competent substrates remains lacking. Here we report a methyl-TROSY based solution NMR study showing that the Escherichia coli version of Hsp70, DnaK, binds to as many as four distinct sites on a small 53-residue client protein, hTRF1. A fraction of hTRF1 chains are also bound to two DnaK molecules simultaneously, resulting in a mixture of DnaK-substrate sub-ensembles that are structurally heterogeneous. The interactions of Hsp70 with a client protein at different sites results in a fuzzy chaperone-substrate ensemble and suggests a mechanism for Hsp70 function whereby the structural heterogeneity of released substrate molecules enables them to circumvent kinetic traps in their conformational free energy landscape and fold efficiently to the native state.
Highlights
The Hsp70 chaperone system plays a vital role in quality control by overseeing the integrity of the cellular proteome in organisms ranging from bacteria to humans (Balchin et al, 2016)
Hsp70 performs a staggering array of functions and its ATP-dependent interaction with client proteins imparts directionality to a number of cellular processes such as protein folding (Hartl et al, 2011), translocation (Kang et al, 1990; Pilon and Schekman, 1999; Mayer, 2013), disaggregation (Nillegoda et al, 2015; Mogk et al, 2015) and oligomer disassembly (Bocking et al, 2011; De Los Rios and Goloubinoff, 2016)
Hsp70 is able to interact with clients having a broad range of conformations (Schlecht et al, 2011; Mashaghi et al, 2016), including unfolded polypeptides emerging from the ribosome (Teter et al, 1999; Deuerling et al, 1999; Willmund et al, 2013), native proteins such as the heat shock transcription factor (Rodriguez et al, 2008), macromolecular assemblies including clathrin-coated vesicles (Sousa et al, 2016), as well as non-native misfolded and aggregated polypeptide chains (Szabo et al, 1994; Mayer and Bukau, 2005; Goloubinoff and De Los Rios, 2007)
Summary
The Hsp chaperone system plays a vital role in quality control by overseeing the integrity of the cellular proteome in organisms ranging from bacteria to humans (Balchin et al, 2016). Hsp performs a staggering array of functions and its ATP-dependent interaction with client proteins imparts directionality to a number of cellular processes such as protein folding (Hartl et al, 2011), translocation (Kang et al, 1990; Pilon and Schekman, 1999; Mayer, 2013), disaggregation (Nillegoda et al, 2015; Mogk et al, 2015) and oligomer disassembly (Bocking et al, 2011; De Los Rios and Goloubinoff, 2016).
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