Molecular Dynamics Simulations Analysis Gaucher Disease

Technical details can be found in our protocol.

A brief check of results

We briefly checked the length of the MD simulations via gmxcheck. Since we used the AGroS paramter -lengthMD 10, we expected the MD simulation to be 10 ns long. gmxcheck reported for each simulation 2001 frames and a time step of 5 ps per frame which amounts to the expected length of 2001*5ps=10ns.

We further investigated the runtime of the MD simulation by looking at the log files. The simulation of the WT took approximately twice as long as the other two mutations since we used only 16 instead of 32 cores. Performing a MD simulation over 1s would take more than 100000 years if the same hardware were used than we did.

<figtable id="tab:runtime">

Name	Runtime	ns/day	years/1s
WT	16h25:59	14.605	187536
L470P	9h20:49	25.677	106699
W209R	9h18:47	25.770	106314

Runtime of the MD simulations and the estimated runtime for a simulation length of 1s. </figtable> The computation of the energy term Coul(T) + LJ [W3-W3] accounted for most (47%) of the floating point operations.

Visualization

For getting a first impression of the MD simulations, we visualized the structures stored in the trajectory files. A course grained animation of the three simulations with 29 frames is shown in <xr id="fig:viz"/>. Altogether, the animations indicate that all three simulations terminated successfully without sever structural changes. Based on the visual inspection, we could not detect any rearrangements which would suggest a malign effect of one of the mutations. However, the active site residue E235 and E340 might be more flexible in L470P compared to the wildtype.

</figure> </figure> </figure>

Wildtype

L470P

W209R

Visualization of the MD simulations. Red: the active site residues E235 and E340; Blue: the mutated residue.

<figure id="fig:gif_wt">

<figure id="fig:gif_L470P">

<figure id="fig:gif_W209R">

Quality assurance

We checked the quality of the MD simulations regarding the convergence of the pressure, temperature, potential energy, and total energy. Strongly fluctuating thermodynamic parameters and insufficiently converging parameters might hint at problems during the MD simulations or a lacking equilibration.

<xr fig="id:qual_pres"/> shows the development of the pressure along the MD simulations. In all three cases, the pressure fluctuates between -200 and 200 bar but the average pressure of 1 bar does not change. In contrast the the wildtype which exhibits a negative drift, the pressure slightly increases in case of L470P and W209R.

</figure> </figure> </figure>

Wildtype (1.0069/-0.0423231)

L470P (0.996977/0.0144563)

W209R (1.00887/0.00535799)

Development of the pressure during the MD simulation. In brackets: average/total-drift.

<figure id="fig:qual_pres_wt">

<figure id="fig:qual_pres_L470P">

<figure id="fig:qual_pres_W209R">

The temperature of 297.9 K remains constant and varies by only +/- 2 K (<xr id="fig:qual_temp"/>).

</figure> </figure> </figure>

Wildtype (297.912/0.00443415)

L470P (297.907/-0.00147577)

W209R (297.915/0.0095005)

Development of the temperature during the MD simulation. In brackets: average/total-drift.

<figure id="fig:qual_temp_wt">

<figure id="fig:qual_temp_L470P">

<figure id="fig:qual_temp_W209R">

Both the potential energy and the total energy reduces at the beginning of the MD simulations. L470P shows the strongest shift towards the negative range, i.e. the stability of the structure increases most.

</figure> </figure> </figure>

Wildtype (-939801/-450.018)

L470P (-940007/-705.14)

W209R (-940568/-355.406)

Development of the potential energy during the MD simulation. In brackets: average/total-drift.

<figure id="fig:qual_pot_wt">

<figure id="fig:qual_pot_L470P">

<figure id="fig:qual_pot_W209R">

</figure> </figure> </figure>

Wildtype (-768901/-447.47)

L470P (-769094/-705.985)

W209R (-769675/-349.952)

Development of the total energy during the MD simulation. In brackets: average/total-drift.

<figure id="fig:qual_tot_wt">

<figure id="fig:qual_tot_L470P">

<figure id="fig:qual_tot_W209R">

Self-interactions between periodic images are unphysical and might disturb the MD simulations. We therefore checked the minimal distance to adjacent cells by taking (a) only C-alpha atoms and (b) also side-chain atoms into account (<xr id="fig:mindist"/>).

In all three simulations, the minimal distance slightly decreases but does not falls below the cut-off distance of 2 nm. If only C-alpha atoms are taking into account, the minimal distance is bout 0.5 nm higher compared to using all protein atoms. This corresponds to an average side-chain length of 5 A. For W209R, the minimal distance drops below 2 nm at 9180 ps but then increases again such that this event does not impact the MD simulation.

</figure> </figure> </figure>

Wildtype (6860: 2.72/3820: 2.08)

L470P (365: 2.28 / 7395: 2.08)

W209R (9720: 2.61 / 9180: 1.95)

Minimal distance to the adjacent cell with respect C-alpha atoms (black) and all protein atoms (red). In brackets: time frame and minimal distance for c-alpha/protein atoms.

<figure id="fig:mindist_wt">

<figure id="fig:mindist_tot_L470P">

<figure id="fig:mindist_tot_W209R">

RMSF

</figure> </figure> </figure>

Wildtype

L470P

W209R

MD simulations with residues colored by the g_rsmf B-factor (rainbow spectrum). Sticks: residues with a B-factor > 120. Purple: the mutated residue.

<figure id="fig:rsmf_wt">

<figure id="fig:rsmf_L470P">

<figure id="fig:rsmf_W209R">

</figure> </figure> </figure>

Wildtype

L470P

W209R

RMSF per residue with respect to the C-alpha atom.

<figure id="fig:rsmf_c_wt">

<figure id="fig:rsmf_c_L470P">

<figure id="fig:rsmf_c_W209R">

</figure> </figure> </figure>

Wildtype

L470P

W209R

RMSF per residue with respect to all atoms.

<figure id="fig:rsmf_p_wt">

<figure id="fig:rsmf_p_L470P">

<figure id="fig:rsmf_p_W209R">

</figure> </figure> </figure>

Wildtype

L470P

W209R

Superposition of the reference structure (blue) and the average g_rmsf (green) structure.

<figure id="fig:rsmf_avg_wt">

<figure id="fig:rsmf_avg_L470P">

<figure id="fig:rsmf_avg_W209R">