Re: [GaMD-discuss] Inquiry regarding 2D PMF reweighting issues with GaMD simulations

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Hi LiaoBing,

There could be a number of things to check and improve:

* Check boost potential statistics in your reweighting output, especially the standard deviation. It needs to be small enough (e.g., <6 kcal/mol) for accurate reweighting.

* User larger cutoff and larger bin sizes; and/or try different reaction coordinates.

* Importantly, it helps to get ~0.5 - 1 million frames in total for accurate reweighting, for which you may extend the simulations, run multiple replicas or save the frames more frequently.

Hope these help,
Yinglong

Yinglong Miao, Ph.D.
Associate Professor
Department of Pharmacology and
Computational Medicine Program
University of North Carolina - Chapel Hill
Tel: 1-919-962-5696
http://miaolab.org<http://miaolab.org/>

Editor-in-Chief
npj Drug Discovery
https://www.nature.com/npjdrugdiscov

On Jan 21, 2026, at 6:20 AM, 廖兵 <202...@zc...> wrote:

You don't often get email from 202...@zc.... Learn why this is important<https://aka.ms/LearnAboutSenderIdentification>

Dear Professor Miao,

Thank you very much for your prompt reply and helpful answers.

I sincerely apologize for the oversight regarding the missing attachment in my previous email. I have re-attached the zip file here, which contains:

  1.  The running scripts and parameter settings.

  2.  The relevant data and log files.

  3.  The resulting figures showing the discontinuity in the FEL.

  4.  The error messages I encountered.

Could you please help me check if the code parameters (such as the bin size and cutoff) are set correctly based on these files? As you suggested, I suspect the discontinuity might be due to a parameter choice issue, but I would value your expert opinion on whether the current configuration causes this or if I should adjust the reaction coordinates.

Thank you again for your time and guidance.

Best regards,

LiaoBing

廖兵
202...@zc...

Original:

  *   From：Miao, Yinglong<Yin...@me...<mailto:Yin...@me...>>
  *   Date：2026-01-21 04:27:04(中国 (GMT+08:00))
  *   To：廖兵<202...@zc...<mailto:202...@zc...>>
  *   Cc：gamd-discuss <gam...@li...<mailto:gam...@li...>>
  *   Subject：Re: [GaMD-discuss] Inquiry regarding 2D PMF reweighting issues with GaMD simulations

Hi LiaoBing,

There was no file attached to your email, but your reweighting procedure sounds good to me. I will try to answer your specific questions below:

  1.  Is my procedure for concatenating weights from multiple log files correct for use with PyReweighting?

Yes

  1.  Does the discontinuity in the FEL suggest an issue with the sampling convergence, or is it likely a parameter issue (e.g., bin size discX/Y=0.3 or cutoff)?

Likely a parameter choice issue. It would help to see your Result Figure. But protein backbone RMSD may not be a good reaction coordinate. One that can reflect different conformations of your system might be preferred.

You may also try using a larger cutoff, for which 10 might be too small.

  1.  Are there specific checks I should perform to verify if the reweighting factors are correctly mapping to the trajectory frames?

You would need match between weights.dat and input_2d.dat frame by frame.

Hope this helps,
Yinglong

Yinglong Miao, Ph.D.
Associate Professor
Department of Pharmacology and
Computational Medicine Program
University of North Carolina - Chapel Hill
Tel: 1-919-962-5696
http://miaolab.org<http://miaolab.org/>

Editor-in-Chief
npj Drug Discovery
https://www.nature.com/npjdrugdiscov

On Jan 19, 2026, at 11:17 AM, 廖兵 <202...@zc...> wrote:

Dear Prof. Miao and the GaMD Team,

I hope this email finds you well.

I am currently conducting Gaussian accelerated Molecular Dynamics (GaMD) simulations on the CB2 receptor system. I am writing to seek your advice regarding some difficulties I have encountered during the reweighting process using PyReweighting.

1. Simulation Context:

I have performed GaMD simulations and have collected data from 10 log files (gamd_GaMD_1.log through gamd_GaMD_10.log) along with their corresponding trajectories.

2. The Workflow:

I followed the standard tutorial to generate the weights and calculate the Free Energy Landscape (FEL).

  *   Step A: Weight Generation

I concatenated the log files and extracted the weights using the following command (ensuring temperature $T=300K$):

Bash

cat gamd_GaMD_{1..10}.log | grep -v "#" | awk '{

    kBT = 0.001987 * 300;

    dV = $7 + $8;

    print dV/kBT " " $2 " " dV
}' > weights.dat

I confirmed that weights.dat has the correct line count matching my combined trajectory frames.

  *   Step B: Reaction Coordinates

I calculated two reaction coordinates (RCs) and saved them into input_2d.dat:

     *   RC1 (X): Protein Backbone RMSD (Residues 0-298).

     *   RC2 (Y): Distance between TM3 and TM6 (Residues 110 and 219).

  *   Step C: PyReweighting Execution

I used PyReweighting-2D.py with the Cumulant Expansion method (amdweight_CE) as follows:

Bash

python PyReweighting-2D.py \

  -cutoff 10 \

  -input input_2d.dat \

  -weight weights.dat \

  -Xdim 0.0 6.0 -discX 0.3 \

  -Ydim 10.0 18.0 -discY 0.3 \

  -Emax 20 \

  -job amdweight_CE \

  > log_reweight_final.txt 2>&1

3. The Issue:

After reweighting, I plotted the FEL using the generated pmf-2D-Final.xvg file (using Python/Matplotlib with tricontourf). However, the resulting landscape appears discontinuous and lacks a clear, defined energy basin.

I am trying to compare my results with a reference study (Nature, Fig 7k, https://www.nature.com/articles/s12276-024-01296-x#Fig7), but my FEL shows significant discrepancies and lacks the expected minimum energy wells.

4. Attachments & Code:

I have attached the plotting script I used and a screenshot of the "strange" FEL I obtained.

  *   Plotting Script: (See below)

  *   Resulting Figure: Final_Landscape_Styled.png (Attached)

  *   Log Files: log_reweight.txt (Attached)

My Plotting Code Snippet:

Python

# (I am using tricontourf to visualize the xvg file)
# ... [Variables setup] ...
cf = ax.tricontourf(x, y, z_relative, levels=levels_range, cmap='jet', extend='max')
# ...

Could you please provide some insights on the following:

  1.  Is my procedure for concatenating weights from multiple log files correct for use with PyReweighting?

  2.  Does the discontinuity in the FEL suggest an issue with the sampling convergence, or is it likely a parameter issue (e.g., bin size discX/Y=0.3 or cutoff)?

  3.  Are there specific checks I should perform to verify if the reweighting factors are correctly mapping to the trajectory frames?

Any guidance or suggestions would be greatly appreciated. Thank you very much for your time and for developing this excellent tool.

Best regards,

LiaoBing

Zhejiang Chinese Medical University

廖兵
202...@zc...

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