# 1 histogram from umbrella sampling?

**URL:** <https://gromacs.bioexcel.eu/t/1-histogram-from-umbrella-sampling/8446>\
**Category:** User discussions\
**Created:** [February 27, 2024, 2:51pm UTC](https://gromacs.bioexcel.eu/t/1-histogram-from-umbrella-sampling/8446 "2024-02-27T14:51:19Z")\
**Posts on this page:** 4\
**Page:** 1

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**Author:** ![oSalad](https://avatars.discourse-cdn.com/v4/letter/o/898d66/32.png) [@oSalad](https://gromacs.bioexcel.eu/u/oSalad)\
**Post date:** [February 27, 2024, 2:51pm UTC](https://gromacs.bioexcel.eu/t/1-histogram-from-umbrella-sampling/8446/1 "2024-02-27T14:51:19Z")

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GROMACS version:  
GROMACS modification: Yes/No  
Here post your question

Hello all,

I recently ran an umbrella simulation, pulling 2 proteins apart. My PMF curve looks correct but for some reasons, I only see 1 peak on my histogram plot. I have changed wham to bins -50 per another post suggestion but that did not seem to help.

 ![Screenshot 2024-02-27 at 8.47.25 AM](https://europe1.discourse-cdn.com/flex017/uploads/bioexcel1/original/2X/7/78fc56049a2062ca9edcd55a7abab2ff85a18b6f.png)

 ![Screenshot 2024-02-27 at 8.47.38 AM](https://europe1.discourse-cdn.com/flex017/uploads/bioexcel1/original/2X/1/1b5749808dc55e65da1ec4e7d6e8641874a53461.png)

Here is my umbrella sampling mdp file (Im 100% sure my steered MD was good):  
title = umbrella sampling md  
define = -DPOSRES\_RECEPTOR  
; Run parameters  
integrator = md  
nsteps = 1000000 ; 2 ns  
dt = 0.002  
; Output control  
nstxout = 5000 ; every 10 ps  
nstvout = 5000  
nstfout = 500  
nstxtcout = 500 ; every 1 ps  
nstenergy = 500

; Bond parameters  
continuation = yes  
constraint\_algorithm = lincs  
constraints = all-bonds ; constrain all bonds  
lincs\_iter = 1  
lincs\_order = 4

; Neighbor searching and vdW  
cutoff-scheme = Verlet  
nstlist = 20  
vdwtype = Cut-off  
vdw-modifier = Potential-shift  
DispCorr = EnerPres  
rvdw = 1.0  
rlist = 1.2  
rcoulomb = 1.0  
coulombtype = PME  
pme\_order = 4  
fourierspacing = 0.125

; Temperature coupling  
tcoupl = v-rescale  
tc-grps = Protein Non-Protein ; two coupling groups  
tau\_t = 1.0 1.0  
ref\_t = 310 310  
; Pressure coupling  
pcoupl = Parrinello-Rahman ; pressure coupling is on for NPT  
pcoupltype = isotropic  
tau\_p = 5.0 ; time constant, in ps  
ref\_p = 1.0 ; reference pressure, in bar  
compressibility = 4.5e-5 ; isothermal compressibility of water, bar^-1  
refcoord\_scaling = com

; Velocity generation  
gen\_vel = no

;COM Pulling  
pull = yes  
pull\_ncoords = 1 ; only one reaction coordinate  
pull\_ngroups = 2 ; two groups defining one reaction coordinate  
pull\_group1\_name = Protein1  
pull\_group2\_name = Protein2  
pull\_coord1\_type = umbrella ; harmonic potential  
pull\_coord1\_geometry = distance ; simple distance increase  
pull\_coord1\_dim = N Y N  
pull\_coord1\_groups = 1 2  
pull\_coord1\_start = yes ; define initial COM distance \> 0  
pull\_coord1\_rate = 0.0 ; restrain in place  
pull\_coord1\_k = 1000 ; kJ mol^-1 nm^-2

Thanks!

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<div class="post-metadata">

**Author:** ![oSalad](https://avatars.discourse-cdn.com/v4/letter/o/898d66/32.png) [@oSalad](https://gromacs.bioexcel.eu/u/oSalad)\
**Post date:** [March 1, 2024, 5:38pm UTC](https://gromacs.bioexcel.eu/t/1-histogram-from-umbrella-sampling/8446/2 "2024-03-01T17:38:59Z")

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My PMF seems to also be more shaky than what I see from other people.

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<div class="post-metadata">

**Author:** ![Stmyo](https://avatars.discourse-cdn.com/v4/letter/s/9fc29f/32.png) [@Stmyo](https://gromacs.bioexcel.eu/u/Stmyo)\
**Post date:** [March 5, 2024, 11:45am UTC](https://gromacs.bioexcel.eu/t/1-histogram-from-umbrella-sampling/8446/3 "2024-03-05T11:45:20Z")

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Hello oSalad,

I had the same issue. Check the histo.xvg file, this can be just an xmgrace problem. I opened the file with excel and the curves looked as they should.

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<div class="post-metadata">

**Author:** ![kush](https://avatars.discourse-cdn.com/v4/letter/k/74df32/32.png) [@kush](https://gromacs.bioexcel.eu/u/kush)\
**Post date:** [February 26, 2025, 6:08am UTC](https://gromacs.bioexcel.eu/t/1-histogram-from-umbrella-sampling/8446/4 "2025-02-26T06:08:02Z")

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You should use this command, xmgrace -nxy histo.xvg