Simulation of the Opening and Closing of Hsp70 Chaperones by Coarse-Grained Molecular Dynamics

Abstract

Heat-shock proteins 70 (Hsp70s) are key molecular chaperones which assist in the folding and refolding/disaggregation of proteins. Hsp70s, which consist of a nucleotide-binding domain (NBD, consisting of NBD-I and NBD-II subdomains) and a substrate-binding domain [SBD, further split into the β-sheet (SBD-β) and α-helical (SBD-α) subdomains], occur in two major conformations having (a) a closed SBD, in which the SBD and NBD domains do not interact, (b) an open SBD, in which SBD-α interacts with NBD-I and SBD-β interacts with the top parts of NBD-I and NBD-II. In the SBD-closed conformation, SBD is bound to a substrate protein, with release occurring after transition to the open conformation. While the transition from the closed to the open conformation is triggered efficiently by binding of adenosine triphosphate (ATP) to the NBD, it also occurs, although less frequently, in the absence of ATP. The reverse transition occurs after ATP hydrolysis. Here, we report canonical and multiplexed replica exchange simulations of the conformational dynamics of Hsp70s using a coarse-grained molecular dynamics approach with the UNRES force field. The simulations were run in the following three modes: (i) with the two halves of the NBD unrestrained relative to each other, (ii) with the two halves of the NBD restrained in an “open” geometry as in the SBD-closed form of DnaK (2KHO), and (iii) the two halves of NBD restrained in a “closed” geometry as in known experimental structures of ATP-bound NBD forms of Hsp70. Open conformations, in which the SBD interacted strongly with the NBD, formed spontaneously during all simulations; the number of transitions was largest in simulations carried out with the “closed” NBD domain, and smallest in those carried out with the “open” NBD domain; this observation is in agreement with the experimentally-observed influence of ATP-binding on the transition of Hsp70’s from the SBD-closed to the SBD-open form. Two kinds of open conformations were observed: one in which SBD-α interacts with NBD-I and SBD-β interacts with the top parts of NBD-I and NBD-II (as observed in the structures of nucleotide exchange factors), and another one in which this interaction pattern is swapped. A third type of motion, in which SBD-α binds to NBD without dissociating from SBD-β was also observed. It was found that the first stage of interdomain communication (approach of SBD-β, to NBD) is coupled with the rotation of the long axes of NBD-I and NBD-II towards each other. To the best of our knowledge, this is the first successful simulation of the full transition of an Hsp70 from the SBD-closed to the SBD-open conformation.

Nanomechanics of the substrate binding domain of Hsp70 determine its allosteric ATP-induced conformational change

“…Single-molecule FRET experiments (22) showed that the open SBD form is present (8-11%) under ADP conditions, supporting our results. A weak interaction between the α/β-folded subdomains and physical separation of subdomains was also observed in native-state molecular simulations by using the replica exchange method or following the dynamics of DnaK over tens of nanoseconds of equilibrium molecular dynamics (23,24). As pointed out above, the subdomain interface is easily perturbed allowing the opening of the lid, whereas β7-β8 are still attached to the β-core, which reduces energy costs.…”

New Strategies for Integrative Dynamic Modeling of Macromolecular Assembly

“…In the case of the barnase-barstar, self-assembled structures were closely resembling the known target complex (Basdevant, Borgis, & Ha-Duong, 2013). Finally, the UNRES force field (Liwo, Czaplewski, Pillardy, & Scheraga, 2001), originally developed to simulate folding and aggregation of small proteins (Czaplewski, Kalinowski, Liwo, & Scheraga, 2009), has been recently used to study the recognition process of Hsp70 chaperones (Gołaś et al, 2012), and the aggregation of the β-amyloid fragments (Aβ 1-28 ), demonstrating that helical intermediate secondary structure elements may play a role during aggregation (Rojas, Liwo, & Scheraga, 2011).…”

Common functionally important motions of the nucleotide‐binding domain of Hsp70

“…3 The SBD-closed (ADP-bound) structure of the DnaK Hsp70 chaperone from E. coli was determined by Zuiderweg and coworkers 10 by NMR spectroscopy (PDB: 2KHO). Recently, 11 we carried out molecular dynamics simulations, using the coarse-grained UNRES force field 12-15 developed in our laboratory, of the conformational transition of DnaK, starting from the SBD-closed 2KHO structure. We determined a probable structure of the SBD-open conformation 11 , which was unknown at the time, and which turned out to be very close to the x-ray structure of the SBD-open (ATP-bound) form of DnaK, which was determined by Mayer and coworkers 3 after the results of our simulations had been published.…”

“…Recently, 11 we carried out molecular dynamics simulations, using the coarse-grained UNRES force field 12-15 developed in our laboratory, of the conformational transition of DnaK, starting from the SBD-closed 2KHO structure. We determined a probable structure of the SBD-open conformation 11 , which was unknown at the time, and which turned out to be very close to the x-ray structure of the SBD-open (ATP-bound) form of DnaK, which was determined by Mayer and coworkers 3 after the results of our simulations had been published. Starting from a homology model of the SBD-open conformation of bovine Hsp70, and using all-atom molecular dynamics with the GROMOS force field, Senet, Ripoll, and coworkers 16 simulated the initial stage of the dissociation of SBD-β from the NBD.…”

“…However, the dynamics of conformational changes, which are critical for understanding the mechanism of allosteric communication (as pointed out by experimental 22 and theoretical 11,16 studies), remains to be explained. From the FRET (Förster resonance energy transfer) study by Mapa et al 25 , it is known that the ADP-bound conformation exhibits a bimodal distance distribution, which suggests conformational transitions between the SBD-open and SBD-closed form; however, FRET cannot provide fine details of the dynamics of conformational changes.…”