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. 2014 Jun 12;10(6):e1003679.
doi: 10.1371/journal.pcbi.1003679. eCollection 2014 Jun.

Computational Modeling of Allosteric Regulation in the hsp90 Chaperones: A Statistical Ensemble Analysis of Protein Structure Networks and Allosteric Communications

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Free PMC article

Computational Modeling of Allosteric Regulation in the hsp90 Chaperones: A Statistical Ensemble Analysis of Protein Structure Networks and Allosteric Communications

Kristin Blacklock et al. PLoS Comput Biol. .
Free PMC article

Abstract

A fundamental role of the Hsp90 chaperone in regulating functional activity of diverse protein clients is essential for the integrity of signaling networks. In this work we have combined biophysical simulations of the Hsp90 crystal structures with the protein structure network analysis to characterize the statistical ensemble of allosteric interaction networks and communication pathways in the Hsp90 chaperones. We have found that principal structurally stable communities could be preserved during dynamic changes in the conformational ensemble. The dominant contribution of the inter-domain rigidity to the interaction networks has emerged as a common factor responsible for the thermodynamic stability of the active chaperone form during the ATPase cycle. Structural stability analysis using force constant profiling of the inter-residue fluctuation distances has identified a network of conserved structurally rigid residues that could serve as global mediating sites of allosteric communication. Mapping of the conformational landscape with the network centrality parameters has demonstrated that stable communities and mediating residues may act concertedly with the shifts in the conformational equilibrium and could describe the majority of functionally significant chaperone residues. The network analysis has revealed a relationship between structural stability, global centrality and functional significance of hotspot residues involved in chaperone regulation. We have found that allosteric interactions in the Hsp90 chaperone may be mediated by modules of structurally stable residues that display high betweenness in the global interaction network. The results of this study have suggested that allosteric interactions in the Hsp90 chaperone may operate via a mechanism that combines rapid and efficient communication by a single optimal pathway of structurally rigid residues and more robust signal transmission using an ensemble of suboptimal multiple communication routes. This may be a universal requirement encoded in protein structures to balance the inherent tension between resilience and efficiency of the residue interaction networks.

Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. The Architecture and Structural Motifs of the Hsp90 Dimers.
The topology and conserved structural regions of the Hsp90 dimer are shown for the crystal structure of yeast ATP-Hsp90 (A), and the crystal structure of ADP-HtpG (B). The structures are shown in a ribbon representation and main structural elements of the Hsp90 dimer are annotated. The Hsp90 domains are colored as follows: Hsp90-NTD (in green), Hsp90-MD (in blue), and Hsp90-CTD (in red). (A) Structural regions of the ATP-Hsp90 dimer. The lid residues 95–123 are highlighted in black sticks. The catalytic loop (371-SEDLPLNLSREMLQQ-385) with a key catalytic residue R-380 is depicted in sticks. The ATP molecules are shown in spheres colored by atoms. The three-helix bundle elements are shown in blue ribbons according to the Hsp90-MD coloration (helix 1: residues 386–408; helix 2: residues 412–431; helix3: residues 435–442) links the inter-domain regions. The inter-domain hinge residues are shown blue spheres. The NTD-MD hinge region (residues 376-LNLSREML-383) also includes catalytic R380. The MD-CTD hinge site includes residues 426-KLGVHE-431. (B) Structural regions of the ADP-HtpG dimer. The functionally important lid region of HtpG (residues 100–126 according to [56]) is highlighted in black sticks as in (A). The ATP molecules are shown in spheres colored by atoms. The three-helix bundle elements are shown in blue ribbons according to the Hsp90-MD coloration (helix 1: residues 336–366 where R336 is the catalytic residue; helix2: residues 368–388; helix 3: residues 393–399). The inter-domain hinge residues are shown in blue spheres. The NTD-MD inter-domain hinge site in HtpG is formed by conserved residues 332-LNVSREIL-339 that contain catalytic R336, and the MD-CTD hinge region includes residues 378-FGLVLKE-384.
Figure 2
Figure 2. Conformational Dynamics of the Hsp90 Crystal Structures.
The computed B-factors are obtained from MD simulations of apo-HtpG (PDB ID 2IOQ) (A); ADP-bound HtpG (PDB ID 2IOP) (B); ATP-bound yeast Hsp90 (PDB ID 2CG9) (C); and ATP-bound Grp94 (PDB ID 2O1U) (D). The NTD residues are shown in green, MD residues are in blue, and CTD residues are in red. The residue-based profiles are based on the consecutive residue numbering adopted from the original crystallographic residue annotation. For clarity, the equilibrium profiles are shown only for one monomer of the homodimer. (A) The fluctuation profile of the apo-HtpG crystal structure has the following residue annotation: NTD (residues 1–219); MD (residues 220–474); CTD (residues 475–577). (B) The profile of the ADP-HtpG crystal structure has the following residue annotation: NTD (residues 1–231); MD (residues 232–486); CTD (residues 487–618). (C) The force constant profile of the yeast ATP-Hsp90 has the following residue annotation: NTD (residues 1–215); MD (residues 216–471); CTD (residues 472–609). (D) The force constant profile of the ATP-Grp94 crystal structure has the following residue annotation: NTD (residues 1–179); MD (residues 180–427); CTD (residues 428–573).
Figure 3
Figure 3. Nucleotide-Dependent Conformational Changes in the HtpG Chaperone.
Conformational changes in the ADP-bound HtpG (A,B) and ATP-bound HtpG complexes (C,D) show that HtpG can experience opening and closing motions that are nucleotide-specific. The normalized frequency of the average distance between the HtpG-NTDs is shown for ADP-HtpG as a green-colored histogram (A) and for ATP-HtpG as red-colored histogram (C). The first bar in both histograms is deliberately truncated to make the small fraction of the distribution corresponding to the NTDs separation visible. The average structure of ADP-HtpG is shown in green ribbons (B) and the average conformation of ATP-HtpG is shown in red ribbons (D). The ADP-bound HtpG structure revealed a partial coordinated separation of the NTDs and MDs. The ATP-bound HtpG complex remained in the closed conformation during simulations.
Figure 4
Figure 4. The Force Constant Stability Analysis of the apo-HtpG Chaperone.
(A) The residue-based force constant profile of apo-HtpG. The NTD residues are in green, MD residues are in blue, and CTD residues are in red. The residue-based dynamic profiles are annotated using the residue numbering in the original crystal structure . The peaks of the force constant profiles corresponding to functionally important residues are indicated by arrows and annotated. Functional residues corresponding to the peaks in the force constant distribution are mapped onto the domain-colored crystal structure of apo-HtpG (B) and onto the functional dynamics profile of apo-HtpG (C). The functional dynamics profile is obtained using PCA of the MD-based conformational ensembles averaged over three lowest frequency modes. The color gradient in from blue to red indicates the decreasing structural stability (or increasing conformational mobility) of protein residues. Functional residues are annotated and shown in spheres, colored according to their domains in (B) and according to the level of rigidity (flexibility) in the functional dynamics profiles (C).
Figure 5
Figure 5. The Force Constant Stability Analysis of the ADP-bound HtpG Chaperone.
(A) The residue-based force constant profile of the ADP-bound HtpG structure. The NTD residues are in green, MD residues are in blue, and CTD residues are in red. The residue-based dynamic profiles are annotated using the residue numbering in the original crystal structure . The peaks of the force constant profiles corresponding to functionally important residues are indicated by arrows and annotated. Functional residues corresponding to the peaks in the force constant distribution are mapped onto the domain-colored crystal structure of ADP-HtpG (B) and onto the functional dynamics profile of ADP-HtpG (C). Functional residues are annotated and shown in spheres and colored as in Figure 4.
Figure 6
Figure 6. The Force Constant Stability Analysis of the ATP-bound yeast Hsp90 Chaperone.
(A) The residue-based force constant profile of the ATP-bound yeast Hsp90 structure. The NTD residues are in green, MD residues are in blue, and CTD residues are in red. The residue-based dynamic profiles are annotated using the residue numbering in the original crystal structure . The peaks of the force constant profiles corresponding to functionally important residues are indicated by arrows and annotated. Functional residues corresponding to the peaks in the force constant distribution are mapped onto the domain-colored crystal structure of the ATP-bound yeast Hsp90 (B) and onto the functional dynamics profile of the ATP-bound yeast Hsp90 (C). Functional residues are annotated and shown in spheres and colored as in Figure 4.
Figure 7
Figure 7. The Network Analysis of the Hsp90 Crystal Structures.
The distribution of hubs (A) and communities (B) in different functional states of Hsp90. The distributions of protein structure network parameters are obtained by averaging computations over MD simulation trajectories. The analysis is based on structurally stable residue interaction networks that remained intact in more than 75% of the simulation snapshots. The distributions are shown for the open solution conformation of HtpG obtained from SAXS studies , (in blue); an apo form of HtpG (PDB ID 2IOQ) (in red); an ADP-bound form of HtpG (PDB ID 2IOP) (in green); an ADP-bound form of the Grp94 homologue (PDB ID 2O1V) (in maroon); an ATP-bound form of the Grp94 homologue (PDB ID 2O1U) (in orange); and an ATP-bound conformation of yeast Hsp90 (PDB ID 2CG9) (in brown).
Figure 8
Figure 8. Structural Maps of Stable Interaction Communities in the Hsp90 Structures.
The distribution of structurally stable interaction communities is shown for the SAXS structure of HtpG (A); the crystal structure of apo HtpG (B); the crystal structure of ADP-HtpG (C); and the crystal structure of yeast ATP-Hsp90 (D). The Hsp90 structures are shown in a ribbon representation and colored according to their domain nomenclature: Hsp90-NTD (in green), Hsp90-MD (in blue), and Hsp90-CTD (in red). The Hsp90 residues that constitute structural communities are shown in spheres and colored according to their respective domains. The residue-based annotation of stable communities is provided and structural positions of communities are indicated by arrows. Structural mapping of stable communities in the solution HtpG structure (A) and ADP-HtpG structure (C) shows a partly disjointed pattern of the interaction networks that are mostly confined within the individual domains. The ATP-bound dimer of yeast Hsp90 has a dense interaction network that includes both the inter-domain and the inter-monomer communities. The Pymol program was used for visualization of the Hsp90 structures (The PyMOL Molecular Graphics System, Version 1.2r3pre, Schrödinger, LLC).
Figure 9
Figure 9. The Degree Distribution of Residue Hubs in the Hsp90 Structures.
The number of hub nodes as a function of the degree of a hub is shown for the solution HtpG structure (in blue); the apo-HtpG structure (in red); the ADP-HtpG structure (in green); the ADP-Grp94 structure (in maroon); the ATP-Grp94 structure (in orange); and the yeast ATP-Hsp90 structure (in brown). The coloring annotation of the distribution is consistent with the annotation of residue hubs in Figure 7.
Figure 10
Figure 10. The Frequency Distributions of the Network Centrality Parameters in Hsp90 Structures.
The frequency distributions of the betweenness values are shown in different functional states of Hsp90: (A) the apo-HtpG structure (red bars); (B) the ADP-HtpG structure (green bars), (C) the yeast ATP-Hsp90 structure (brown bars), and (D) the ATP-Grp94 structure (orange bars). The residue-based betweenness distributions point to small-world organization of the interaction networks in the Hsp90 structures.
Figure 11
Figure 11. The Residue-Based Betweenness Profiles of the HtpG Structures.
Dynamics-based analysis of network centrality in the HtpG crystal structures. The residue-based betweenness profiles are shown for the apo HtpG crystal structure (A) and the ADP-HtpG crystal structure (C). The betweenness profiles are shown in green for the NTD residues, in blue for the MD residues, and in red for the CTD residues. The peaks of the betweenness profiles corresponding to functionally important residues are indicated by arrows and annotated. Structural mapping of high betweenness residues that correspond to functionally important sites is shown for the apo HtpG crystal structure (B) and the ADP-HtpG crystal structure (D). The protein structures are shown in a backbone trace representation and domain-colored: NTD (in green), MD (in blue), and CTD (in red). The functional residues of high centrality are shown in spheres and colored according to their respective domains. Structural positions of high centrality functional residues are indicated by arrows.
Figure 12
Figure 12. The Residue-Based Betweenness Profiles of the ATP-bound yeast Hsp90 and Grp94 Structures.
The residue-based betweenness profiles are shown for the crystal structure of yeast ATP-Hsp90 (A) and the crystal structure of ATP-Grp94 (C). The betweenness profiles are shown in green for the NTD residues, in blue for the MD residues, and in red for the CTD residues. The peaks of the betweenness profiles corresponding to functionally important residues are indicated by arrows and annotated. Structural mapping of high betweenness residues that correspond to functionally important sites is shown for the crystal structure of yeast ATP-Hsp90 (B) and the crystal structure of ATP-Grp94 (D). The protein structures are shown in a backbone trace protein representation and colored according to their domain nomenclature as in Figure 11. The functional residues of high centrality are shown in spheres and colored according to their respective domains. Structural positions of high centrality residues are indicated by arrows.

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This work is supported by funding from Chapman University. No additional external funding received for this study. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
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