GROMACS Wizard – Step 4: NPT Equilibration

This section is part of the GROMACS Wizard tutorial.

Once the system has been successfully minimized and the system’s temperature has stabilized at the desired value, it is necessary to apply pressure to the system until it reaches the correct density. This second equilibration phase is aimed at stabilizing the system’s density at the desired value by performing equilibration using the NPT ensemble (constant Number of particles, Pressure, and Temperature) also known as “isothermal-isobaric”.

Switch to the Equilibrate (NPT) tab.

Selecting input structure

When launching the NPT Equilibration step, GROMACS Wizard requires you to provide one of the following:

  • The path to a GRO file resulting from the previous step: either a GRO file resulting from the Minimization stepNVT Equilibration step, or the previous launch of an equilibration step.
  • The path to a batch project, that has been minimized and/or equilibrated using NVT ensemble or resulted from one of the other steps.

If you want to proceed from the previous step you can simply click on the Auto-fill button highlighted in the image below. This will automatically fill the path based on the previous project, whether a GRO file or a batch project from the previous successful run (e.g. from the NVT Equilibration).

You can also choose the input GRO file yourself by clicking on the “” button.

Choosing parameters

GROMACS molecular dynamics parameters for NPT equilibration are present in the Parameters section of the NPT Equilibration tab. By default, these parameters are populated with default values that are suitable for usual energy minimization runs. You can modify these parameters as needed. In the Parameters section, you will find the parameters that are most likely to be changed often, like the integration time step and the number of steps.

The timeframe for such a procedure depends upon the size and contents of the system, but in NPT, the running average of the density of the system should reach a plateau at the desired value. Typically, 100 ps should be a good start. Please note that pressure is a quantity that might fluctuate widely throughout equilibration or simulation. If the density has not yet stabilized (i.e. the pressure-related terms are slow to converge) in the given timeframe, additional time will be required – you can simply run the NPT equilibration step again by providing the input data from the previous NPT equilibration step.

The other GROMACS molecular dynamics parameters can be accessed by clicking on the “All…” button highlighted in the image above. To learn more on how to apply custom parameters please check the Applying custom parameters section.

Note: The position restraint (POSRES*) options are filled automatically based on the input system.

In the “Pressure coupling” section, you can modify the corresponding parameters, e.g. barostat, time constant, and the reference pressure. For most projects, you can use the exponential relaxation pressure coupling with time constant including a stochastic term (c-rescale) and the time constant of about 5 ps, as shown in the image below.

Note: Please ensure that the temperature coupling parameters correspond to the ones from the NVT equilibration, and the temperature in the velocity generation (if it is on) as well.

For the sake of this tutorial, reset the parameters to their default values.

To restore the parameters to their default values click on the “Reset” button in the Advanced parameters window. To load the parameters from an MDP file from some other project click the “Load from file…” button. To save the parameters in an MDP file click the “Save as…” button.

Note: the modified parameters are saved on closing SAMSON, so the next time you will have the previously saved parameters.

Run NPT Equilibration

GROMACS Wizard gives you several possibilities:

  • Generate inputs – generates a ready-to-use project that you can, for example, run on your local cluster.
  • Equilibrate locally – launches the computations locally on your PC using the shipped GROMACS or the one you specified in the settings (see Using custom GROMACS version and performance parameters).
  • Equilibrate in the cloud – launches the computations in the cloud. This is suitable for big systems that would otherwise require too many resources to do it locally. This will prompt you to select the Cloud machine, etc. Note that launching in the Cloud requires computing credits. See Launching Computations in the Cloud for more information.

Now, simply click on the Equilibrate locally button to launch the calculations on your PC. Some pop-ups might appear informing you about the current steps or possible warnings/issues if there are any.

Depending on your PC, the equilibration of the system in the tutorial might take a few minutes. You can see the current progress in the Output window that should pop up.

During these calculations, you can still use SAMSON and GROMACS Wizard thanks to the job manager of the GROMACS Wizard Extension. You can always access the list of the local GROMACS jobs and their state via the “Local jobs” button.

Importing the results

If you launched a single project and there are no other projects in the local job queue then after the computations are done a pop-up will appear asking for import options. If you launched multiple jobs or a batch job then you can check their state in the Local jobs. You can choose whether to import the whole trajectory, only the last frame, or some range of frames, what type of Periodic Boundary Condition treatment to apply, and on what to center the system. For example, as shown in the image below, you can choose to center the system on the Protein.

If you do not want to import the trajectory you can simply click Cancel – this will not delete any results and will still generate the plot.

You can access the results in the results folder specified at the top of the GROMACS Wizard. The folders with results are named with the launch date and time, and the step description (for the NPT Equilibration step, the folder suffix is “_npt“).


You can check the plots describing the evolution of the system’s pressure and density over simulation time at the bottom of the tab in the Plots section.

In this example, this plot demonstrates that the density is stabilized at 1030 kg/m3 which is close to the experimental value of 1000 kg/m3 and the expected density of the SPC/E model is about 1008 kg/m3. We can see that the density values are stable over time, indicating that the system is well-equilibrated now with respect to pressure and density.

The plots are automatically generated and saved when the job is finished and the results are loaded. If you would like to save the plot, click on the Save button on top of the figure.

Checking results

It is important to check whether the system has reached the desired density and whether that density stabilized. Please note, that density might fluctuate around the desired value. If the system has not reached the desired density or has not yet stabilized at it then you would need to launch additional NPT Equilibration starting from the results of this NPT Equilibration. For that, set the input GRO file to the current NPT Equilibration results (you can simply click on the Auto-fill button to set it to the latest results).

Once the system’s density has stabilized at the desired value, we will proceed to the actual simulation.


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