Breaking the Deadlock of Molecular Chaperones

Protein folding in the cell requires ATP-driven chaperone machines. It is poorly understood, however, how these machines fold proteins. Here we propose that the conserved Hsp70 and Hsp90 chaperones support formation of the folding nucleus by providing a gradient of decreasing hydrophobicity. Early on the folding pathway Hsp70 uses its highly hydrophobic binding pocket to recover a stalled, unproductive folding intermediate. The aggressive nature of Hsp70 action, however, blocks productive folding by grabbing hydrophobic, core-forming segments. This precludes on-pathway nucleation at high, physiological Hsp70 levels. Transfer to the less hydrophobic Hsp90 enables the intermediate to resume forming its folding nucleus. Subsequently, the protein enters a spontaneous folding trajectory towards its native state, independent of the ATPase activities of both Hsp70 and Hsp90. Our findings provide a general mechanistic concept for chaperoned protein folding. Introduction It is the primary sequence of the protein that determines its native fold . Proteins condense around an initial nucleus to ultimately shape up to always the same structure . In the cell, conserved families of molecular chaperones support folding of their proteins in an energyconsuming manner, presumably by repeated cycles of binding and release . The molecular determinants of assisted protein folding, however, remain largely enigmatic. Here we describe that the ubiquitous, ATP-dependent chaperone machines Hsp70 and Hsp90 form a conserved cascade that promotes spontaneous protein folding by a stop-start mechanism. Hsp90 acts downstream of Hsp70, but its contribution to protein folding is unclear . Under certain conditions Hsp70 can refold proteins in the absence of Hsp90 . However, molecular understanding of the Hsp70 folding mechanism requires solving two paradoxes: (i) Hsp70 binds peer-reviewed) is the author/funder. All rights reserved. No reuse allowed without permission. The copyright holder for this preprint (which was not . http://dx.doi.org/10.1101/176693 doi: bioRxiv preprint first posted online Aug. 15, 2017;