The force-dependent mechanism of DnaK-mediated mechanical folding

Science Advances
Judit Perales-CalvoSergi Garcia-Manyes

Abstract

It is well established that chaperones modulate the protein folding free-energy landscape. However, the molecular determinants underlying chaperone-mediated mechanical folding remain largely elusive, primarily because the force-extended unfolded conformation fundamentally differs from that characterized in biochemistry experiments. We use single-molecule force-clamp spectroscopy, combined with molecular dynamics simulations, to study the effect that the Hsp70 system has on the mechanical folding of three mechanically stiff model proteins. Our results demonstrate that, when working independently, DnaJ (Hsp40) and DnaK (Hsp70) work as holdases, blocking refolding by binding to distinct substrate conformations. Whereas DnaK binds to molten globule-like forms, DnaJ recognizes a cryptic sequence in the extended state in an unanticipated force-dependent manner. By contrast, the synergetic coupling of the Hsp70 system exhibits a marked foldase behavior. Our results offer unprecedented molecular and kinetic insights into the mechanisms by which mechanical force finely regulates chaperone binding, directly affecting protein elasticity.

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Citations

Jul 3, 2019·International Journal of Molecular Sciences·Le Minh BuiSun Chang Kim
Apr 8, 2020·Cell Stress & Chaperones·Dhawal ChoudharyCiro Cecconi
Apr 8, 2020·Cell Stress & Chaperones·Miranda P Collier, Justin L P Benesch
Sep 16, 2020·Chemical Society Reviews·Marc MoraSergi Garcia-Manyes
Apr 30, 2020·Nature Communications·Jaime Andrés Rivas-PardoJorge Alegre-Cebollada
Mar 26, 2021·Nano Letters·Andrew StannardSergi Garcia-Manyes
Aug 22, 2021·Journal of Molecular Biology·Rafayel PetrosyanMichael T Woodside

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Methods Mentioned

BETA
nucleotide exchange
protein folding
atomic force spectroscopy
atomic force microscopy
pull-down
gel filtration
electrophoresis

Software Mentioned

NAMD
Igor Pro
Python
SciPy
WaveMetrics

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