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representative motion corrected cryo em micrograph  (Thermo Fisher)


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    Thermo Fisher representative motion corrected cryo em micrograph
    (A) Schematic diagram showing the domain organization of the proteins present in the RRHCC complex. The dashed and non-colored sections indicate regions where there is no interpretable density. <t>(B)</t> <t>Cryo-EM</t> map of the RRHCC complex (RRHCC_a6) at 2.6 Å global resolution. The map is colored according to each subunit as in (A). (C) Cartoon representation of the RRHCC assembly in complex with ATP (sphere representation). The N-lobe sections of RAF1.1 and RAF1.2 associated with HSP90-CDC37 are depicted as thicker coils. The black oval highlights the interaction of RAF1.2 N-lobe with CDC37.1-MD and HSP90-A. (D) Zoom of the N-lobe of RAF1.2 showing an almost completely folded αC helix, which is captured between the CTD of HSP90-A and CDC37.1-MD. (E) Two perpendicular views of the interface formed by the RAF1.2 αC helix, CDC37.1-MD, and HSP90-A (green, orange, and blue, respectively) in the local map RRHCC_Cloc. Key interacting residues are labeled. The accompanying cartoon of αC in the mold illustrates the arrangement. The residues at the base of the αC helix set its position and register by engaging distinct hydrophobic (hp), basic, and acidic patches. (F) Surface representation of CDC37.1-MD (model from RRHCC_Cloc map), colored by electrostatic potential. Two opposite views are shown: CDC37.1-MD region (upper panel) and the corresponding HSP90-A interface (lower panel). RAF1.2 (green ribbon) fits into the central groove of CDC37.1 MD, which features two oppositely charged patches at its exit (upper panel). RAF1.2 residue H402 inserts into a pocket of HSP90-A (lower panel). A fully extended αC helix from the folded RAF1 structure (PDB: 8CPD, transparent grey) would clash with the acidic patch of CDC37.1-MD. Relevant RAF1.2 residues are labeled. (G) Same as in (F) with the αC of RAF1.2 displayed with its electrostatic potential surface, showing its complementation with the electrostatic potential of CDC37 and HSP90. Pairs of interacting residues between CDC37.1-MD and HSP90-A are indicated. (H) The αC helix of RAF1.2 is substituted with its fully folded conformation from PDB entry 8CPD to illustrate that the folded helix is incompatible with the acidic patch located at the exit of CDC37.1-MD. (I) Electrostatic potential surface of the folded RAF1 N-lobe, with the αC helix shown as a grey ribbon. For comparison, the fitted RAF1.2 model from the RRHCC_Cloc map is displayed in green in the same orientation as in (A). (J) Interface between the RAF1 N-lobes in a dimer (PDB:8CPD), with residues that contact CDC37-MD highlighted. One subunit is colored according to the scheme used in . (K) Schematic illustrating how the secondary structure elements of the RAF1 N-lobe assemble into the final folded state. When the αC helix is released from the CDC37-MD, the segment of RAF1 that is clamped by HSP90 must be released upon HSP90 opening. Then, together with the final 30 N-terminal residues in the RRHCC, the kinase can fold the final two β-sheets. These β-sheets are inserted between the αC helix and the remaining β-strands building the RAF1 N-lobe. (L) Comparison of the model from the local map and the folded RAF1 N-lobe (PDB: 8CPD), aligned at the αC helix region. The inset shows the fully folded RAF1 structure, with the N-lobe in grey and the C-lobe in white. Also see Figure S2, SI1-4, and Table S2.
    Representative Motion Corrected Cryo Em Micrograph, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "The HSP90–CDC37 Chaperone System Orchestrates RAF1 Kinase Activation Through a Pre-Dimerization Mechanism"

    Article Title: The HSP90–CDC37 Chaperone System Orchestrates RAF1 Kinase Activation Through a Pre-Dimerization Mechanism

    Journal: bioRxiv

    doi: 10.64898/2026.03.25.713956

    (A) Schematic diagram showing the domain organization of the proteins present in the RRHCC complex. The dashed and non-colored sections indicate regions where there is no interpretable density. (B) Cryo-EM map of the RRHCC complex (RRHCC_a6) at 2.6 Å global resolution. The map is colored according to each subunit as in (A). (C) Cartoon representation of the RRHCC assembly in complex with ATP (sphere representation). The N-lobe sections of RAF1.1 and RAF1.2 associated with HSP90-CDC37 are depicted as thicker coils. The black oval highlights the interaction of RAF1.2 N-lobe with CDC37.1-MD and HSP90-A. (D) Zoom of the N-lobe of RAF1.2 showing an almost completely folded αC helix, which is captured between the CTD of HSP90-A and CDC37.1-MD. (E) Two perpendicular views of the interface formed by the RAF1.2 αC helix, CDC37.1-MD, and HSP90-A (green, orange, and blue, respectively) in the local map RRHCC_Cloc. Key interacting residues are labeled. The accompanying cartoon of αC in the mold illustrates the arrangement. The residues at the base of the αC helix set its position and register by engaging distinct hydrophobic (hp), basic, and acidic patches. (F) Surface representation of CDC37.1-MD (model from RRHCC_Cloc map), colored by electrostatic potential. Two opposite views are shown: CDC37.1-MD region (upper panel) and the corresponding HSP90-A interface (lower panel). RAF1.2 (green ribbon) fits into the central groove of CDC37.1 MD, which features two oppositely charged patches at its exit (upper panel). RAF1.2 residue H402 inserts into a pocket of HSP90-A (lower panel). A fully extended αC helix from the folded RAF1 structure (PDB: 8CPD, transparent grey) would clash with the acidic patch of CDC37.1-MD. Relevant RAF1.2 residues are labeled. (G) Same as in (F) with the αC of RAF1.2 displayed with its electrostatic potential surface, showing its complementation with the electrostatic potential of CDC37 and HSP90. Pairs of interacting residues between CDC37.1-MD and HSP90-A are indicated. (H) The αC helix of RAF1.2 is substituted with its fully folded conformation from PDB entry 8CPD to illustrate that the folded helix is incompatible with the acidic patch located at the exit of CDC37.1-MD. (I) Electrostatic potential surface of the folded RAF1 N-lobe, with the αC helix shown as a grey ribbon. For comparison, the fitted RAF1.2 model from the RRHCC_Cloc map is displayed in green in the same orientation as in (A). (J) Interface between the RAF1 N-lobes in a dimer (PDB:8CPD), with residues that contact CDC37-MD highlighted. One subunit is colored according to the scheme used in . (K) Schematic illustrating how the secondary structure elements of the RAF1 N-lobe assemble into the final folded state. When the αC helix is released from the CDC37-MD, the segment of RAF1 that is clamped by HSP90 must be released upon HSP90 opening. Then, together with the final 30 N-terminal residues in the RRHCC, the kinase can fold the final two β-sheets. These β-sheets are inserted between the αC helix and the remaining β-strands building the RAF1 N-lobe. (L) Comparison of the model from the local map and the folded RAF1 N-lobe (PDB: 8CPD), aligned at the αC helix region. The inset shows the fully folded RAF1 structure, with the N-lobe in grey and the C-lobe in white. Also see Figure S2, SI1-4, and Table S2.
    Figure Legend Snippet: (A) Schematic diagram showing the domain organization of the proteins present in the RRHCC complex. The dashed and non-colored sections indicate regions where there is no interpretable density. (B) Cryo-EM map of the RRHCC complex (RRHCC_a6) at 2.6 Å global resolution. The map is colored according to each subunit as in (A). (C) Cartoon representation of the RRHCC assembly in complex with ATP (sphere representation). The N-lobe sections of RAF1.1 and RAF1.2 associated with HSP90-CDC37 are depicted as thicker coils. The black oval highlights the interaction of RAF1.2 N-lobe with CDC37.1-MD and HSP90-A. (D) Zoom of the N-lobe of RAF1.2 showing an almost completely folded αC helix, which is captured between the CTD of HSP90-A and CDC37.1-MD. (E) Two perpendicular views of the interface formed by the RAF1.2 αC helix, CDC37.1-MD, and HSP90-A (green, orange, and blue, respectively) in the local map RRHCC_Cloc. Key interacting residues are labeled. The accompanying cartoon of αC in the mold illustrates the arrangement. The residues at the base of the αC helix set its position and register by engaging distinct hydrophobic (hp), basic, and acidic patches. (F) Surface representation of CDC37.1-MD (model from RRHCC_Cloc map), colored by electrostatic potential. Two opposite views are shown: CDC37.1-MD region (upper panel) and the corresponding HSP90-A interface (lower panel). RAF1.2 (green ribbon) fits into the central groove of CDC37.1 MD, which features two oppositely charged patches at its exit (upper panel). RAF1.2 residue H402 inserts into a pocket of HSP90-A (lower panel). A fully extended αC helix from the folded RAF1 structure (PDB: 8CPD, transparent grey) would clash with the acidic patch of CDC37.1-MD. Relevant RAF1.2 residues are labeled. (G) Same as in (F) with the αC of RAF1.2 displayed with its electrostatic potential surface, showing its complementation with the electrostatic potential of CDC37 and HSP90. Pairs of interacting residues between CDC37.1-MD and HSP90-A are indicated. (H) The αC helix of RAF1.2 is substituted with its fully folded conformation from PDB entry 8CPD to illustrate that the folded helix is incompatible with the acidic patch located at the exit of CDC37.1-MD. (I) Electrostatic potential surface of the folded RAF1 N-lobe, with the αC helix shown as a grey ribbon. For comparison, the fitted RAF1.2 model from the RRHCC_Cloc map is displayed in green in the same orientation as in (A). (J) Interface between the RAF1 N-lobes in a dimer (PDB:8CPD), with residues that contact CDC37-MD highlighted. One subunit is colored according to the scheme used in . (K) Schematic illustrating how the secondary structure elements of the RAF1 N-lobe assemble into the final folded state. When the αC helix is released from the CDC37-MD, the segment of RAF1 that is clamped by HSP90 must be released upon HSP90 opening. Then, together with the final 30 N-terminal residues in the RRHCC, the kinase can fold the final two β-sheets. These β-sheets are inserted between the αC helix and the remaining β-strands building the RAF1 N-lobe. (L) Comparison of the model from the local map and the folded RAF1 N-lobe (PDB: 8CPD), aligned at the αC helix region. The inset shows the fully folded RAF1 structure, with the N-lobe in grey and the C-lobe in white. Also see Figure S2, SI1-4, and Table S2.

    Techniques Used: Cryo-EM Sample Prep, Labeling, Residue, Comparison

    Composition of the HSP90 ATP-binding sites in the different cryo-EM maps (see also SI4). Each panel shows the density for a given site as a transparent surface, together with the modeled ligands at that site (phosphate is labeled as PO4). The central schematic summarizes the ATP-hydrolysis trajectory along the RHC and RRHCC assemblies, starting from an open HSP90 dimer. The upper branch corresponds to the RHC pathway, and the lower branch to the RRHCC classes. Blue arrows indicate increased HSP90 ATPase activity, red arrows indicate decreased activity, and dashed lines denote alternative transitions between the RHC and RRHCC pathways.
    Figure Legend Snippet: Composition of the HSP90 ATP-binding sites in the different cryo-EM maps (see also SI4). Each panel shows the density for a given site as a transparent surface, together with the modeled ligands at that site (phosphate is labeled as PO4). The central schematic summarizes the ATP-hydrolysis trajectory along the RHC and RRHCC assemblies, starting from an open HSP90 dimer. The upper branch corresponds to the RHC pathway, and the lower branch to the RRHCC classes. Blue arrows indicate increased HSP90 ATPase activity, red arrows indicate decreased activity, and dashed lines denote alternative transitions between the RHC and RRHCC pathways.

    Techniques Used: Binding Assay, Cryo-EM Sample Prep, Labeling, Activity Assay

    (A) Two views of the cryo-EM density map of class RHCp23_c with the domain organization of p23. (B) Ribbon representation of RHCp23_c in two orientations matching (a). (C) Cryo-EM map of class RHCp23_p. (D) Side-by-side view of the complexes of GR and RAF1 with HSP90-CDC37 showing the C-lobe of the RAF1 kinase domain in the opposite site of the GR complex while the MD of CDC37 is positioned in the same area. Detailed view of the superimposition of the complexes depicting the location of the P23 protein. (E) Comparison of CDC37-MD orientation in RHC_c1 and RHCp23_c. Ribbon diagrams of both structures in three perpendicular views; CDC37-MD movement is indicated by black arrows. The dashed line on top of the p23 helical section indicates the proposed modeling. (F) Corresponding cryo-EM maps of these regions, colored by protein. Additional density on CDC37-MD is shown in magenta. (G) Model for p23-assisted release of RAF1 from CDC37-MD in RHC. p23 first binds at the HSP90 dimer interface (1), enabling its flexible tail to contact CDC37-MD (2). This interaction tilts CDC37-MD (3), promoting release of the captured RAF1 N-lobe (5). Alternatively, the p23 tail may compete with the partially unfolded RAF1 N-lobe and block its binding (4). After ATP hydrolysis, the HSP90 dimer opens and releases RAF1 (6).
    Figure Legend Snippet: (A) Two views of the cryo-EM density map of class RHCp23_c with the domain organization of p23. (B) Ribbon representation of RHCp23_c in two orientations matching (a). (C) Cryo-EM map of class RHCp23_p. (D) Side-by-side view of the complexes of GR and RAF1 with HSP90-CDC37 showing the C-lobe of the RAF1 kinase domain in the opposite site of the GR complex while the MD of CDC37 is positioned in the same area. Detailed view of the superimposition of the complexes depicting the location of the P23 protein. (E) Comparison of CDC37-MD orientation in RHC_c1 and RHCp23_c. Ribbon diagrams of both structures in three perpendicular views; CDC37-MD movement is indicated by black arrows. The dashed line on top of the p23 helical section indicates the proposed modeling. (F) Corresponding cryo-EM maps of these regions, colored by protein. Additional density on CDC37-MD is shown in magenta. (G) Model for p23-assisted release of RAF1 from CDC37-MD in RHC. p23 first binds at the HSP90 dimer interface (1), enabling its flexible tail to contact CDC37-MD (2). This interaction tilts CDC37-MD (3), promoting release of the captured RAF1 N-lobe (5). Alternatively, the p23 tail may compete with the partially unfolded RAF1 N-lobe and block its binding (4). After ATP hydrolysis, the HSP90 dimer opens and releases RAF1 (6).

    Techniques Used: Cryo-EM Sample Prep, Comparison, Blocking Assay, Binding Assay



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    representative motion corrected cryo em micrograph  (Thermo Fisher)
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    Thermo Fisher representative motion corrected cryo em micrograph
    (A) Schematic diagram showing the domain organization of the proteins present in the RRHCC complex. The dashed and non-colored sections indicate regions where there is no interpretable density. <t>(B)</t> <t>Cryo-EM</t> map of the RRHCC complex (RRHCC_a6) at 2.6 Å global resolution. The map is colored according to each subunit as in (A). (C) Cartoon representation of the RRHCC assembly in complex with ATP (sphere representation). The N-lobe sections of RAF1.1 and RAF1.2 associated with HSP90-CDC37 are depicted as thicker coils. The black oval highlights the interaction of RAF1.2 N-lobe with CDC37.1-MD and HSP90-A. (D) Zoom of the N-lobe of RAF1.2 showing an almost completely folded αC helix, which is captured between the CTD of HSP90-A and CDC37.1-MD. (E) Two perpendicular views of the interface formed by the RAF1.2 αC helix, CDC37.1-MD, and HSP90-A (green, orange, and blue, respectively) in the local map RRHCC_Cloc. Key interacting residues are labeled. The accompanying cartoon of αC in the mold illustrates the arrangement. The residues at the base of the αC helix set its position and register by engaging distinct hydrophobic (hp), basic, and acidic patches. (F) Surface representation of CDC37.1-MD (model from RRHCC_Cloc map), colored by electrostatic potential. Two opposite views are shown: CDC37.1-MD region (upper panel) and the corresponding HSP90-A interface (lower panel). RAF1.2 (green ribbon) fits into the central groove of CDC37.1 MD, which features two oppositely charged patches at its exit (upper panel). RAF1.2 residue H402 inserts into a pocket of HSP90-A (lower panel). A fully extended αC helix from the folded RAF1 structure (PDB: 8CPD, transparent grey) would clash with the acidic patch of CDC37.1-MD. Relevant RAF1.2 residues are labeled. (G) Same as in (F) with the αC of RAF1.2 displayed with its electrostatic potential surface, showing its complementation with the electrostatic potential of CDC37 and HSP90. Pairs of interacting residues between CDC37.1-MD and HSP90-A are indicated. (H) The αC helix of RAF1.2 is substituted with its fully folded conformation from PDB entry 8CPD to illustrate that the folded helix is incompatible with the acidic patch located at the exit of CDC37.1-MD. (I) Electrostatic potential surface of the folded RAF1 N-lobe, with the αC helix shown as a grey ribbon. For comparison, the fitted RAF1.2 model from the RRHCC_Cloc map is displayed in green in the same orientation as in (A). (J) Interface between the RAF1 N-lobes in a dimer (PDB:8CPD), with residues that contact CDC37-MD highlighted. One subunit is colored according to the scheme used in . (K) Schematic illustrating how the secondary structure elements of the RAF1 N-lobe assemble into the final folded state. When the αC helix is released from the CDC37-MD, the segment of RAF1 that is clamped by HSP90 must be released upon HSP90 opening. Then, together with the final 30 N-terminal residues in the RRHCC, the kinase can fold the final two β-sheets. These β-sheets are inserted between the αC helix and the remaining β-strands building the RAF1 N-lobe. (L) Comparison of the model from the local map and the folded RAF1 N-lobe (PDB: 8CPD), aligned at the αC helix region. The inset shows the fully folded RAF1 structure, with the N-lobe in grey and the C-lobe in white. Also see Figure S2, SI1-4, and Table S2.
    Representative Motion Corrected Cryo Em Micrograph, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/representative motion corrected cryo em micrograph/product/Thermo Fisher
    Average 99 stars, based on 1 article reviews
    representative motion corrected cryo em micrograph - by Bioz Stars, 2026-05
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    (A) Schematic diagram showing the domain organization of the proteins present in the RRHCC complex. The dashed and non-colored sections indicate regions where there is no interpretable density. (B) Cryo-EM map of the RRHCC complex (RRHCC_a6) at 2.6 Å global resolution. The map is colored according to each subunit as in (A). (C) Cartoon representation of the RRHCC assembly in complex with ATP (sphere representation). The N-lobe sections of RAF1.1 and RAF1.2 associated with HSP90-CDC37 are depicted as thicker coils. The black oval highlights the interaction of RAF1.2 N-lobe with CDC37.1-MD and HSP90-A. (D) Zoom of the N-lobe of RAF1.2 showing an almost completely folded αC helix, which is captured between the CTD of HSP90-A and CDC37.1-MD. (E) Two perpendicular views of the interface formed by the RAF1.2 αC helix, CDC37.1-MD, and HSP90-A (green, orange, and blue, respectively) in the local map RRHCC_Cloc. Key interacting residues are labeled. The accompanying cartoon of αC in the mold illustrates the arrangement. The residues at the base of the αC helix set its position and register by engaging distinct hydrophobic (hp), basic, and acidic patches. (F) Surface representation of CDC37.1-MD (model from RRHCC_Cloc map), colored by electrostatic potential. Two opposite views are shown: CDC37.1-MD region (upper panel) and the corresponding HSP90-A interface (lower panel). RAF1.2 (green ribbon) fits into the central groove of CDC37.1 MD, which features two oppositely charged patches at its exit (upper panel). RAF1.2 residue H402 inserts into a pocket of HSP90-A (lower panel). A fully extended αC helix from the folded RAF1 structure (PDB: 8CPD, transparent grey) would clash with the acidic patch of CDC37.1-MD. Relevant RAF1.2 residues are labeled. (G) Same as in (F) with the αC of RAF1.2 displayed with its electrostatic potential surface, showing its complementation with the electrostatic potential of CDC37 and HSP90. Pairs of interacting residues between CDC37.1-MD and HSP90-A are indicated. (H) The αC helix of RAF1.2 is substituted with its fully folded conformation from PDB entry 8CPD to illustrate that the folded helix is incompatible with the acidic patch located at the exit of CDC37.1-MD. (I) Electrostatic potential surface of the folded RAF1 N-lobe, with the αC helix shown as a grey ribbon. For comparison, the fitted RAF1.2 model from the RRHCC_Cloc map is displayed in green in the same orientation as in (A). (J) Interface between the RAF1 N-lobes in a dimer (PDB:8CPD), with residues that contact CDC37-MD highlighted. One subunit is colored according to the scheme used in . (K) Schematic illustrating how the secondary structure elements of the RAF1 N-lobe assemble into the final folded state. When the αC helix is released from the CDC37-MD, the segment of RAF1 that is clamped by HSP90 must be released upon HSP90 opening. Then, together with the final 30 N-terminal residues in the RRHCC, the kinase can fold the final two β-sheets. These β-sheets are inserted between the αC helix and the remaining β-strands building the RAF1 N-lobe. (L) Comparison of the model from the local map and the folded RAF1 N-lobe (PDB: 8CPD), aligned at the αC helix region. The inset shows the fully folded RAF1 structure, with the N-lobe in grey and the C-lobe in white. Also see Figure S2, SI1-4, and Table S2.

    Journal: bioRxiv

    Article Title: The HSP90–CDC37 Chaperone System Orchestrates RAF1 Kinase Activation Through a Pre-Dimerization Mechanism

    doi: 10.64898/2026.03.25.713956

    Figure Lengend Snippet: (A) Schematic diagram showing the domain organization of the proteins present in the RRHCC complex. The dashed and non-colored sections indicate regions where there is no interpretable density. (B) Cryo-EM map of the RRHCC complex (RRHCC_a6) at 2.6 Å global resolution. The map is colored according to each subunit as in (A). (C) Cartoon representation of the RRHCC assembly in complex with ATP (sphere representation). The N-lobe sections of RAF1.1 and RAF1.2 associated with HSP90-CDC37 are depicted as thicker coils. The black oval highlights the interaction of RAF1.2 N-lobe with CDC37.1-MD and HSP90-A. (D) Zoom of the N-lobe of RAF1.2 showing an almost completely folded αC helix, which is captured between the CTD of HSP90-A and CDC37.1-MD. (E) Two perpendicular views of the interface formed by the RAF1.2 αC helix, CDC37.1-MD, and HSP90-A (green, orange, and blue, respectively) in the local map RRHCC_Cloc. Key interacting residues are labeled. The accompanying cartoon of αC in the mold illustrates the arrangement. The residues at the base of the αC helix set its position and register by engaging distinct hydrophobic (hp), basic, and acidic patches. (F) Surface representation of CDC37.1-MD (model from RRHCC_Cloc map), colored by electrostatic potential. Two opposite views are shown: CDC37.1-MD region (upper panel) and the corresponding HSP90-A interface (lower panel). RAF1.2 (green ribbon) fits into the central groove of CDC37.1 MD, which features two oppositely charged patches at its exit (upper panel). RAF1.2 residue H402 inserts into a pocket of HSP90-A (lower panel). A fully extended αC helix from the folded RAF1 structure (PDB: 8CPD, transparent grey) would clash with the acidic patch of CDC37.1-MD. Relevant RAF1.2 residues are labeled. (G) Same as in (F) with the αC of RAF1.2 displayed with its electrostatic potential surface, showing its complementation with the electrostatic potential of CDC37 and HSP90. Pairs of interacting residues between CDC37.1-MD and HSP90-A are indicated. (H) The αC helix of RAF1.2 is substituted with its fully folded conformation from PDB entry 8CPD to illustrate that the folded helix is incompatible with the acidic patch located at the exit of CDC37.1-MD. (I) Electrostatic potential surface of the folded RAF1 N-lobe, with the αC helix shown as a grey ribbon. For comparison, the fitted RAF1.2 model from the RRHCC_Cloc map is displayed in green in the same orientation as in (A). (J) Interface between the RAF1 N-lobes in a dimer (PDB:8CPD), with residues that contact CDC37-MD highlighted. One subunit is colored according to the scheme used in . (K) Schematic illustrating how the secondary structure elements of the RAF1 N-lobe assemble into the final folded state. When the αC helix is released from the CDC37-MD, the segment of RAF1 that is clamped by HSP90 must be released upon HSP90 opening. Then, together with the final 30 N-terminal residues in the RRHCC, the kinase can fold the final two β-sheets. These β-sheets are inserted between the αC helix and the remaining β-strands building the RAF1 N-lobe. (L) Comparison of the model from the local map and the folded RAF1 N-lobe (PDB: 8CPD), aligned at the αC helix region. The inset shows the fully folded RAF1 structure, with the N-lobe in grey and the C-lobe in white. Also see Figure S2, SI1-4, and Table S2.

    Article Snippet: Cryo-EM data and processing workflow, related to , , 3, 4, Table S2. (A) Representative motion-corrected cryo-EM micrograph of the RHC particles in vitreous ice imaged with a Titan Krios Falcon 4i direct detector camera (-2.3 μm defocus). (B) Characteristic reference-free 2D averages of the RHC particles.

    Techniques: Cryo-EM Sample Prep, Labeling, Residue, Comparison

    Composition of the HSP90 ATP-binding sites in the different cryo-EM maps (see also SI4). Each panel shows the density for a given site as a transparent surface, together with the modeled ligands at that site (phosphate is labeled as PO4). The central schematic summarizes the ATP-hydrolysis trajectory along the RHC and RRHCC assemblies, starting from an open HSP90 dimer. The upper branch corresponds to the RHC pathway, and the lower branch to the RRHCC classes. Blue arrows indicate increased HSP90 ATPase activity, red arrows indicate decreased activity, and dashed lines denote alternative transitions between the RHC and RRHCC pathways.

    Journal: bioRxiv

    Article Title: The HSP90–CDC37 Chaperone System Orchestrates RAF1 Kinase Activation Through a Pre-Dimerization Mechanism

    doi: 10.64898/2026.03.25.713956

    Figure Lengend Snippet: Composition of the HSP90 ATP-binding sites in the different cryo-EM maps (see also SI4). Each panel shows the density for a given site as a transparent surface, together with the modeled ligands at that site (phosphate is labeled as PO4). The central schematic summarizes the ATP-hydrolysis trajectory along the RHC and RRHCC assemblies, starting from an open HSP90 dimer. The upper branch corresponds to the RHC pathway, and the lower branch to the RRHCC classes. Blue arrows indicate increased HSP90 ATPase activity, red arrows indicate decreased activity, and dashed lines denote alternative transitions between the RHC and RRHCC pathways.

    Article Snippet: Cryo-EM data and processing workflow, related to , , 3, 4, Table S2. (A) Representative motion-corrected cryo-EM micrograph of the RHC particles in vitreous ice imaged with a Titan Krios Falcon 4i direct detector camera (-2.3 μm defocus). (B) Characteristic reference-free 2D averages of the RHC particles.

    Techniques: Binding Assay, Cryo-EM Sample Prep, Labeling, Activity Assay

    (A) Two views of the cryo-EM density map of class RHCp23_c with the domain organization of p23. (B) Ribbon representation of RHCp23_c in two orientations matching (a). (C) Cryo-EM map of class RHCp23_p. (D) Side-by-side view of the complexes of GR and RAF1 with HSP90-CDC37 showing the C-lobe of the RAF1 kinase domain in the opposite site of the GR complex while the MD of CDC37 is positioned in the same area. Detailed view of the superimposition of the complexes depicting the location of the P23 protein. (E) Comparison of CDC37-MD orientation in RHC_c1 and RHCp23_c. Ribbon diagrams of both structures in three perpendicular views; CDC37-MD movement is indicated by black arrows. The dashed line on top of the p23 helical section indicates the proposed modeling. (F) Corresponding cryo-EM maps of these regions, colored by protein. Additional density on CDC37-MD is shown in magenta. (G) Model for p23-assisted release of RAF1 from CDC37-MD in RHC. p23 first binds at the HSP90 dimer interface (1), enabling its flexible tail to contact CDC37-MD (2). This interaction tilts CDC37-MD (3), promoting release of the captured RAF1 N-lobe (5). Alternatively, the p23 tail may compete with the partially unfolded RAF1 N-lobe and block its binding (4). After ATP hydrolysis, the HSP90 dimer opens and releases RAF1 (6).

    Journal: bioRxiv

    Article Title: The HSP90–CDC37 Chaperone System Orchestrates RAF1 Kinase Activation Through a Pre-Dimerization Mechanism

    doi: 10.64898/2026.03.25.713956

    Figure Lengend Snippet: (A) Two views of the cryo-EM density map of class RHCp23_c with the domain organization of p23. (B) Ribbon representation of RHCp23_c in two orientations matching (a). (C) Cryo-EM map of class RHCp23_p. (D) Side-by-side view of the complexes of GR and RAF1 with HSP90-CDC37 showing the C-lobe of the RAF1 kinase domain in the opposite site of the GR complex while the MD of CDC37 is positioned in the same area. Detailed view of the superimposition of the complexes depicting the location of the P23 protein. (E) Comparison of CDC37-MD orientation in RHC_c1 and RHCp23_c. Ribbon diagrams of both structures in three perpendicular views; CDC37-MD movement is indicated by black arrows. The dashed line on top of the p23 helical section indicates the proposed modeling. (F) Corresponding cryo-EM maps of these regions, colored by protein. Additional density on CDC37-MD is shown in magenta. (G) Model for p23-assisted release of RAF1 from CDC37-MD in RHC. p23 first binds at the HSP90 dimer interface (1), enabling its flexible tail to contact CDC37-MD (2). This interaction tilts CDC37-MD (3), promoting release of the captured RAF1 N-lobe (5). Alternatively, the p23 tail may compete with the partially unfolded RAF1 N-lobe and block its binding (4). After ATP hydrolysis, the HSP90 dimer opens and releases RAF1 (6).

    Article Snippet: Cryo-EM data and processing workflow, related to , , 3, 4, Table S2. (A) Representative motion-corrected cryo-EM micrograph of the RHC particles in vitreous ice imaged with a Titan Krios Falcon 4i direct detector camera (-2.3 μm defocus). (B) Characteristic reference-free 2D averages of the RHC particles.

    Techniques: Cryo-EM Sample Prep, Comparison, Blocking Assay, Binding Assay