# Please help me understand these values from the tutorial **URL:** \ **Category:** pmx\ **Created:** [November 10, 2023, 10:16am UTC](https://ask.bioexcel.eu/t/please-help-me-understand-these-values-from-the-tutorial/4664 "2023-11-10T10:16:24Z")\ **Posts on this page:** 5\ **Page:** 1
**Author:** ![mstieffe](https://avatars.discourse-cdn.com/v4/letter/m/a698b9/32.png) [@mstieffe](https://ask.bioexcel.eu/u/mstieffe)\ **Post date:** [November 10, 2023, 10:16am UTC](https://ask.bioexcel.eu/t/please-help-me-understand-these-values-from-the-tutorial/4664/1 "2023-11-10T10:16:24Z")
Hello, I am having a hard time understanding the dG and ddG values that I obtain when going through the [amino acid mutation tutorial](http://pmx.mpibpc.mpg.de/summerSchool2020_tutorial1/index.html). - 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 ---
**Author:** ![bgroot](https://avatars.discourse-cdn.com/v4/letter/b/dc4da7/32.png) [@bgroot](https://ask.bioexcel.eu/u/bgroot)\ **Post date:** [November 10, 2023, 12:05pm UTC](https://ask.bioexcel.eu/t/please-help-me-understand-these-values-from-the-tutorial/4664/2 "2023-11-10T12:05:19Z")
Hi, > [@mstieffe](#): > > - 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? 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 > [@mstieffe](#): > > - 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? I think you should look up S2A instead of A2S from the tripeptide database? best Bert ---
**Author:** ![mstieffe](https://avatars.discourse-cdn.com/v4/letter/m/a698b9/32.png) [@mstieffe](https://ask.bioexcel.eu/u/mstieffe)\ **Post date:** [November 10, 2023, 12:33pm UTC](https://ask.bioexcel.eu/t/please-help-me-understand-these-values-from-the-tutorial/4664/3 "2023-11-10T12:33:46Z")
Dear Bert, 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? 🙂 Best, Marc ---
**Author:** ![vgapsys](https://avatars.discourse-cdn.com/v4/letter/v/7993a0/32.png) [@vgapsys](https://ask.bioexcel.eu/u/vgapsys)\ **Post date:** [November 22, 2023, 8:59am UTC](https://ask.bioexcel.eu/t/please-help-me-understand-these-values-from-the-tutorial/4664/4 "2023-11-22T08:59:01Z")
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 ---
**Author:** ![mstieffe](https://avatars.discourse-cdn.com/v4/letter/m/a698b9/32.png) [@mstieffe](https://ask.bioexcel.eu/u/mstieffe)\ **Post date:** [November 23, 2023, 2:27pm UTC](https://ask.bioexcel.eu/t/please-help-me-understand-these-values-from-the-tutorial/4664/5 "2023-11-23T14:27:27Z")
thanks for the explanation!