# UMBRELLA SAMPILNG pull reference

**URL:** <https://gromacs.bioexcel.eu/t/umbrella-sampilng-pull-reference/12947>\
**Category:** User discussions\
**Tags:** umbrella-sampling\
**Created:** [November 19, 2025, 9:22am UTC](https://gromacs.bioexcel.eu/t/umbrella-sampilng-pull-reference/12947 "2025-11-19T09:22:45Z")\
**Posts on this page:** 2\
**Page:** 1

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**Author:** ![sankalp1999](https://avatars.discourse-cdn.com/v4/letter/s/439d5e/32.png) [@sankalp1999](https://gromacs.bioexcel.eu/u/sankalp1999)\
**Post date:** [November 19, 2025, 9:22am UTC](https://gromacs.bioexcel.eu/t/umbrella-sampilng-pull-reference/12947/1 "2025-11-19T09:22:45Z")

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GROMACS version:2022.5  
GROMACS modification: Yes/No  
I have performed the umbrella sampling simulations by pulling the protein away from the membrane.  
pull = yes  
pull\_ncoords = 1 ; only one reaction coordinate  
pull\_ngroups = 2 ; two groups defining one reaction coordinate  
pull\_group1\_name = PROTEIN  
pull\_group2\_name = ECLIP  
pull-pbc-ref-prev-step-com = yes  
pull\_coord1\_type = umbrella ; harmonic potential  
pull\_coord1\_geometry = distance ; simple distance increase  
pull\_coord1\_dim = N N Y  
pull\_coord1\_groups = 1 2  
pull\_coord1\_start = yes ; define initial COM distance \> 0  
pull\_coord1\_rate = 0.005 ; 0.005 nm per ps  
pull\_coord1\_k = 5000 ; kJ mol^-1 nm^-2

I got the PMF, but it did not show the potential well. To get the well, I want to pull it towards the membrane. Initially, my reference point was the protein COM and the membrane upper leaflet headgroup COM. When I pull the protein towards the membrane by setting  
**pull\_coord1\_rate = -0.005 ; 0.005 nm per ps**

I get the error

**Pull reference distance for coordinate 1 (-0.000041) needs to be non-negative**

I suppose it’s because the protein is trying to cross the reference and, hence, getting a negative distance. Should I change the reference to the lower leaflet COM and pull it?  
Will it cause an error in the WHAM calculation because changing the reference will make the distances inconsistent?

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<div class="post-metadata">

**Author:** ![hess](https://dub1.discourse-cdn.com/flex017/user_avatar/gromacs.bioexcel.eu/hess/32/416_2.png) [@hess](https://gromacs.bioexcel.eu/u/hess)\
**Post date:** [November 19, 2025, 9:28am UTC](https://gromacs.bioexcel.eu/t/umbrella-sampilng-pull-reference/12947/2 "2025-11-19T09:28:32Z")

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You should have used geometry=direction, not distance. My guess is that you can change to direction for the points close to zero and reuse all you current data with gmx wham.

Why are you using pull-pbc-ref-prev-step-com = yes? This should not be necessary.