Fig. 4: Phosphorylation of key residues located in the charged linker of HSP90 leads to a conformational shift in the linker, exposing the middle ___domain of the protein.

a Model of the HSP90–HSP90–HSP70–HSP70–HOP assembly used for the molecular dynamics simulations. A and B, protomers A and B, respectively. b Protein secondary structure elements (SSE) like alpha-helices and beta-strands of the charged linker of protomer A of ATP-bound HSP90 monitored throughout the molecular dynamics simulation. WT (HSP90^S226/S255), phosphomimetic (HSP90^S226E/S255E), and non-phosphorylatable (HSP90^S226A/S255A) mutants were analyzed. Each pentameric assembly was simulated three times for 100 ns, yielding similar results across simulations. The plot on the left reports SSE distribution by residue index throughout the charged linker and the plot on the right monitors each residue and its SSE assignment over time. Schematic illustrating the primary structure of the full-length HSP90 with color-coded domains is also shown: NTD N-terminal ___domain; MD middle ___domain and CTD C-terminal ___domain. The charged linker (CL) and the ___location of the two key serine residues are also shown (top inset). The gray bar indicates the CL segment encompassing residues 218–232. c Cartoon representation of ATP-bound HSP90 protomer A in assemblies with either the phosphomimetic (HSP90^S226E/S255E) or non-phosphorylatable (HSP90^S226A/S255A) mutants. The figure depicts the reference trajectory and representative trajectories from n = 1000 simulation frames. The inset illustrates the surfaces available for the interaction between HSP90 A and HSP70 A when the CL is in the up conformation. The arrow indicates the ___location of the key beta-strand in the charged linker. See also Supplementary Figs. 5–9.