Hello,
I am having a hard time understanding the dG and ddG values that I obtain when going through the amino acid mutation tutorial.

I downloaded the dhdl files for the slow growth. Is this slow growth approach what is referred to as equilibrium thermodynamic integration (TI) in the literature? When I use gmx analyze -f forward_dhdl.xvg -integrate i obtain a value for dG=375964.46875 kJ/mol. This does not sound reasonable to me, what am i missing here?

I also downloaded the data for the fast growth and used analyze_dhdl.py -fA eqA/morphes/frame*/dgdl.xvg -fB eqB/morphes/frame*/dgdl.xvg -t 300 --nbins 25 to analyze it. I obtain a value for dG_folded=39.05 kJ/mol. The value for A2S mutation in the tripeptide from the website gives dG_unfolded=-39.15 kJ/mol. That would mean ddG=dG_folded-dG_unfolded=78.2kJ/mol. This is quite different from the provided experimental value of ddG=1.8kJ/mol. Am i misinterpreting this?

Any help to understand those discrepancies is very much appreciated. Thanks in advance!
Best,
Marc

gmx analyze just integrates. In this case, the x-axis is time (not lambda), which runs from 0 to 10000 (and not from 0 to 1 as lambda does). That way, you should divide the obtained value by 10000 to get a deltaG. This would be 37.6 kJ/mol in this case

I think you should look up S2A instead of A2S from the tripeptide database?

Thank you so much for your insightful response. It has been incredibly helpful.

You’re absolutely right; I should have used the S2A tripeptide instead of A2S. I apologize for that oversight. After making this correction, I obtain ddG=-0.1 ± 0.22 kJ/mol, which is still not within error for the experimental value of ddg=-1.8kJ/mol ( i expect its minus 1.8kJ/mol as it is stated to be mildly stabilizing mutation). But i guess this is as accurate as it gets? Or are there any more mistakes I made on the way?

The computed ddG seems fine. Several things to keep in mind:

experimental error. Although in the tutorial (and in the paper) it is not explicitly provided, it should not be ignored that the experimental ddG has an associated uncertainty;

alchemical methods on average yield accuracy of ~1 kcal/mol, so the calculated ddG will not always match the experimental value exactly