Binding pocket dynamics along the recovery stroke of human β-cardiac myosin

Figures

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  Fig 1. Modelling the recovery stroke. A/B. Close-up view of the OM binding site in PR (A) and PPS (B) structures of cardiac myosin (cartoon) in complex with OM (spheres). The following colour code was used for the different sub-domains: N-terminal domain: orange, L50K: grey, U50K: white, relay helix: magenta, converter and lever arm (CLD): blue. C/D. The atoms used to define the collective variables D1 (C) and A1 (D) are represented as grey spheres and connected with dotted lines. The CLD is shown as coloured cartoon in the PR (cyan) and PPS (pink) conformation, while the rest of the protein is shown in white. E. Time evolution of the CLD RMSD from the reference PR (4P7H*, cyan) and PPS (5N6A*, pink) conformations during the 200-ns apoPR-to-PPS SMD simulation. The RMSD was calculated over the Cα atoms of the CLD domain after a best-fit superimposition of the Cα atoms of the rest of the protein. F. Snapshots extracted from the 200-ns apoPR-to-PPS SMD simulation in panel E, showing the rotation of the CLD domain (blue) and the formation of the kink in the relay helix (magenta). G. Close-up view of the SH1 and relay helices at the end of the 200-ns PR-to-PPS SMD trajectory (red),…

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  Fig 2. Change of shape and volume of the binding site along the recovery stroke. A. Isosurfaces connecting points with 30% pocket frequency as determined by MDpocket in 5 representative batches (B1, B7, B10, B15 and B20) from the 200-ns PR-to-PPS SMD trajectory. Three types of pockets were detected near the CLD (blue): a top pocket (light blue) in B1 and B10, a bottom pocket (green) in B10, and a top+bottom pocket where the two subpockets are joined together (cyan) in B7, B15 and B20. B. Surface representation of the binding region for representative structures of five batches along the 200-ns PR-to-PPS SMD simulation. The atoms lining the pockets are coloured according to their sub-domain: orange for the N-terminal domain, grey for L50K, magenta for the relay helix and blue for the converter and lever arm. Atoms from residues that are found to be part of a pocket in all the representative batches are highlighted as cyan spheres. The OM structure in the PR and PPS binding modes is superimposed to the B1 and B20 frames, respectively, and shown as sticks. C. Box plots of pocket volumes for twenty batches along the 200-ns PR-to-PPS SMD trajectory. For batches where the top and bottom…

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  Fig 3. Change of binding site polarity along the recovery stroke. A/B. Box plots of the normalized polarity (A) and hydrophobicity (B) scores as determined by MDpocket for twenty batches of the 200-ns PR-to-PPS SMD trajectory. The polarity score increases during the simulation from ~11 to ~ 28, while the hydrophobicity one decreases from ~38 to ~12. C. Surface representation of the binding region for five representative batches coloured according to the residue type. The atoms lining the pockets are shown in yellow (hydrophobic residues) or purple (polar residues). The same structures and pockets represented in Fig 2B are used. The OM structure in the PR and PPS binding modes is superimposed to the B1 and B20 frames, respectively, and shown as sticks.

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  Fig 4. Intermediates along the recovery stroke. A/B. Free energy landscapes along the CVs with labelling of the minima a, b, d and e. Close-up views of the PR basin from the PR-to-PPS (A) and of the PPS basin from the PPS-to-PR (B) US calculations are shown. C. Scatterplot of the CV coordinates measured on X-ray structures and the a-e minima from US energy landscapes. 5N69 (chain B) [13]: bovine β-cardiac myosin (OM-bound), 5N6A (chain A) [13]: bovine β-cardiac myosin (OM-free), 5O2L (chain A) [29]: myosin VI (recovery stroke intermediate), 4P7H (chain A) [12]: human β-cardiac myosin (OM-free), 4PA0 (chain A) [12]: human β-cardiac myosin (OM-bound). For 5N6A, the structure with gaps completed and energy-minimised as described in reference [14] was used (5N6A*) since 5N6A does not contain one of the amino acids needed for the CV calculation (M776). For 5O2L, D1 was calculated as R136-F763 Cα-atom distance, while A1 was calculated as the angle formed by Q441, I457 and I769 Cα atoms. D. Cartoon representation of intermediate structures b (left), c (middle) and d (right) superimposed to the X-ray structures (CLD only) closest to them. The CLD is highlighted in colour.