Figure 1. Global structural changes depending on box size.

Panel A: Overall structure of the $\alpha 1\beta 1\alpha 2\beta 2$ hemoglobin tetramer. The His146 side chains (green spheres) are specifically indicated. Panel B: Temporal change of the C$\alpha$–C$\alpha$ distance between His146${}_{\beta 1}$ and His146${}_{\beta 2}$. The dashed lines (black, orange, red) indicate transition points for the 75, 90, and 120 box, respectively. Cyan and blue arrows indicate the values of the corresponding observable found for the deoxy T${}_0$ (2DN2) and oxy R${}_4$ (2DN3) states, respectively.

Figure 1—figure supplement 1. Tetrameric hemoglobin solvated in boxes of different sizes.

Tetrameric hemoglobin solvated in boxes of size: (from left to right) 75, 90, 120 and 150 Å. Spheres correspond to Na${}^{+}$ and Cl${}^{-}$ ions, added to neutralize the system and to achieve a biologically appropriate salt concentration of 0.15 mol/L.

Figure 1—figure supplement 2. Global structural changes depending on box size.

Panel A: Overall structure of the $\alpha 1\beta 1\alpha 2\beta 2$ hemoglobin tetramer. The iron atoms (Fe, yellow spheres) and His146 side chains (green spheres) are specifically indicated. The angle $\theta$ measures the angle between the two planes containing His146${}_{\beta 1}$Fe${}_{\beta 1}$Fe${}_{\alpha 1}$ and His146${}_{\beta 2}$Fe${}_{\beta 2}$Fe${}_{\alpha 2}$, respectively. Panel B: temporal change of (from top to bottom) the C$\alpha$–C$\alpha$ distance between His146${}_{\beta 1}$ and His146${}_{\beta 2}$, the C$\alpha$–C$\alpha$ distance between His143${}_{\beta 1}$ and His143${}_{\beta 2}$, C$\alpha$ RMSD relative to the 2DN2 X-ray structure, and the angle $\theta$. The dashed lines (black, orange, red) indicate transition points for the 75, 90, and 120 Å boxes, respectively. Cyan and blue arrows indicate the values of the corresponding observables found for the deoxy T${}_0$ (2DN2) and oxy R${}_4$ (2DN3) states, respectively. Top panel is also given in the main MS.

Figure 1—figure supplement 3. Local structural rearrangement of the C-terminus of the $\beta$ chain.

From top to bottom: the Ramachandran $\varphi$-angle of His143, Lys144, Tyr145 and His146. Left panels report time series for $\beta 1$, and right panels those for $\beta 2$. Cyan and blue arrows indicate the corresponding values found in crystal structure of the deoxy (2DN2) and oxy (2DN3) state, respectively. Color code as in Figure 1—figure supplement 1.

Figure 1—figure supplement 4. Local structural changes around His146.

Left: Interactions involving His146${}_{\beta }$. Right: Interactions involving His146${}_{\beta }$. From top to bottom: [a] water-mediated contact between (His146${}_{\beta }$)COO${}^{-}$–OC(Pro37${}_{\alpha }$), [b] the salt bridge between (His146${}_{\beta }$)COO${}^{-}$–NZ(Lys40${}_{\alpha }$), [c] the contact between (His146${}_{\beta }$)COO${}^{-}$–NZ(Lys132${}_{\beta }$), [d] the contact between (His146${}_{\beta }$)COO${}^{-}$ and NE(His2${}_{\beta }$) and [e] the salt bridge between (His146${}_{\beta }$)NE2 and COO${}^{-}$(Asp94${}_{\beta }$). Cyan and blue arrows indicate the corresponding values found in crystal structure of the deoxy (2DN2) and oxy (2DN3) state, respectively. The dashed lines indicate breaking points along the 1$\mu$s MD simulation and the black arrows indicates the first to break. The straight green line represents the averaged distance over 1 $\mu$s of MD simulation for the 150 Å box as reference.

Figure 1—figure supplement 5. Structural transition details.

(A) Close-up of the [600;650] and [440;520] ns interval of Figure 1—figure supplement 1, illustrating the order of events for transitions in the 120 (left panel) and 90 Å box (right panel), respectively. Black arrows indicates the transmission of the structural modifications. (B) Structural details for every quantity from the 90 Å box at 10 and 800 ns, respectively. The dashed black arrow indicates the separation distance between $\beta 1$ and $\beta 2$.

Figure 1—figure supplement 6. Temporal structural changes of the C$\alpha$–C$\alpha$ distance between His146${}_{\beta 1}$ and His146${}_{\beta 2}$ in the 75 Å box for different simulation conditions.

Temporal structural changes of the C$\alpha$–C$\alpha$ distance between His146${}_{\beta 1}$ and His146${}_{\beta 2}$, depending on the protonation state of His 146 in the 75 Å box with and without scaled protein-water interactions.