Dear Carlos,
Thank you so much for your prompt reply clarifying these two points.
Sincerely yours,
Ruth
> On 3 May 2019, at 17:14, Carlos Simmerling <carlos.simmerling.gmail.com> wrote:
>
> I don't think it matters much if the r1 and r4 are constant or vary, as
> long as they are far away from any values sampled in that window. It's
> important, as you say, that the umbrella potential is harmonic so the r1
> and r4 should be far enough away that the MD during that window does not
> encounter regions with the linear potential.
>
> For distances, the rad-->deg factor is not used, but you should always
> carefully check the conversion of the force constant between the version
> using k(r-r0)^2 and 1/2 k(r-r0)^2.
> for example, see this helpful description in the amber archive:
>
> From: Brian Radak <brian.radak.accts.gmail.com
> <brian.radak.accts.gmail.com?Subject=Re%3A%20%5BAMBER%5D%20Umbrella%20Sampling%20and%20wham>
>>
> Date: Sat, 30 May 2015 13:13:37 -0500
>
> Most WHAM programs assume a restraint pre-factor of 1/2. The AMBER
> nmropt module, however, does not. Therefore you should multiply most
> force constants by 2 in the metafile.
>
> Also, angle and dihedral force constants are specified in
> kcal/mol-rad^2, while the WHAM code usually assumes it is in the same
> units as the coordinate values (i.e. degrees). Therefore you also need
> to convert those force constants to kcal/mol-deg^2.
>
> Cheers,
> Brian
>
> On Fri, May 3, 2019 at 11:10 AM Ruth Helena Tichauer <rhtichau.laas.fr>
> wrote:
>
>> Dear all,
>>
>> I’m also a beginner in Umbrella Sampling methods and the WHAM.
>>
>> Differently from Daniel, I’m using as the reaction coordinate a linear
>> combination of distances but as him, I’ve run the various windows changing
>> the r1 and r4 values. Thus, I’m strikingly surprised to learn that these
>> values should remain constant for all windows although it also sounds
>> logic/coherent. Nevertheless, I’m even more surprised given that in the
>> tutorial (http://ambermd.org/tutorials/advanced/tutorial17/section2.htm)
>> it is stated that WHAM code of Alan Grossfield expects a harmonic potential
>> and so, in order to have a perfectly harmonic potential, this values are
>> equally distant from the r2=r3 value i.e. if r2=r3=30 and r1=-30, then r4
>> is set to r4=90, subsequently, if the next value of r2=r3=40, r1 and r4 are
>> updated to r1=-20 and r4=100, and so on..
>>
>> So now, the two following questions arise for me:
>>
>> 1. Is the WHAM used in the Amber tutorial (that of Alan Grossfield) still
>> valid when the values of r1 and r4 are maintained constant throughout the
>> umbrella sampling simulations?
>> 2. Concerning the conversion factor for getting the force constant in the
>> correct units, can the WHAM be used when using distance reaction
>> coordinates, and in particular linear combination of distances, rather than
>> some angle or dihedral? I’d like that this point be clarified (again) as
>> the conversion factor indicated in the amber tutorial contains a rad -> deg
>> factor.
>>
>> Thank you for any insight on this matter,
>>
>> Sincerely yours,
>>
>> Ruth
>>
>>> On 18 Apr 2019, at 21:35, Zachary Fallon <zachary.fallon.stonybrook.edu>
>> wrote:
>>>
>>>
>>> Hi Daniel,
>>> Your PMF indeed looks very periodic, which is usually a symptom of
>> some
>>> underlying problem with your umbrella sampling or WHAM inputs. I have
>>> produced similar (incorrect) PMFs with that kind of periodicity in two
>>> circumstances, once was when I ran umbrella sampling with varying r1
>> and r4
>>> values across the different windows. I couldn't tell from your last
>> email --
>>> do these runs you're showing here use different r1 and r4 values
>> across the
>>> windows, or did you change them to be constant? And when running WHAM,
>> you
>>> didn't by chance turn on periodicity?Â
>>> The other time I got PMFs like that was when my WHAM inputs were
>> wrong. Are
>>> your data files separate for each window? Try plotting each window
>>> separately. And just in case, when running WHAM each of your windows
>> should
>>> have its own file containing the distances throughout that window's
>>> trajectory, not a single file containing the distances for every
>> window.Â
>>> Along those lines, is there any chance your metadata file has wrong
>> target
>>> values or force constants for any windows?
>>> Also, try looking at orthogonal degrees of freedom other than the
>> distance
>>> you're running umbrella sampling on. Maybe take a look at peptide
>> end-to-end
>>> distances, rmsd to the initial conformation, phi/psi values for a
>> specific
>>> dihedral, etc. You want to make sure that other orthogonal
>> characteristics
>>> of your system overlap with adjacent windows, not just the coordinate
>> you're
>>> running umbrella sampling on. Seems like 5 ns would not be long enough
>> to
>>> sample a 20-residue peptide at each window, but I lack the context of
>> your
>>> system.
>>> Finally, I'm attaching a Python script below that you can use to plot
>>> histograms. It takes as input 60 x-y space-delimited column files, as
>> well
>>> as a name for your output file.
>>> Hope this helps,
>>> --Zachary
>>>
>>> On Thu, Apr 18, 2019 at 2:28 PM Daniel Fernández Remacha
>>> <[1]dnlfr1994.gmail.com> wrote:
>>>
>>> Hi Zachary
>>> The peptide is about 20 residues long and we are using an alpha carbon
>> of
>>> one of the residues of the peptide to separate it from the CoM of
>> several
>>> alpha carbons in the protein.
>>> By the density of the points we are getting with 5ns production, I
>> would say
>>> that sampling is enough. Let me show you a picture of a small part of
>> this
>>> umbrella. (XmGrace representation in "segment" setting with Dot width
>> of
>>> 0.5)
>>> image.png
>>> What do you think? We guessed from this, that overlap between windows
>> and
>>> sampling are OK; maybe a bit more sampling would be better, but I
>> think this
>>> should do. However, I would still like to check histogram overlaying,
>> as
>>> shown in the tutorial. I am having problems to obtain this plot
>> though. Do
>>> you know of any script that could do this easily for about 60 windows?
>>> Also, this is the PMF that we get with 64 windows. Just in case it
>> gives a
>>> better idea of what the problem may be.
>>> image.png
>>> Thank you very much for your help!! Really appreciated!Â
>>> Daniel Fernández
>>> El mié., 17 abr. 2019 a las 19:22, Zachary Fallon
>>> (<[2]zachary.fallon.stonybrook.edu>) escribió:
>>>
>>> Hi Daniel,
>>> I was actually just talking to someone about a system similar to yours
>> the
>>> other day -- specifically pulling a peptide out of a protein pocket.
>> Using
>>> umbrella sampling for this is tricky because, if I recall my
>> conversation
>>> correctly, not only do you have to sample each distance sufficiently,
>> but
>>> you also have to sample every conformation of the peptide at each
>> distance.
>>> Depending on the size of your peptide this may be challenging to
>>> insurmountable, I believe we concluded that using something like
>> accelerated
>>> MD might be better for this application, but you may have to consult
>> the
>>> literature to see what others have done in similar systems. Other
>> users can
>>> correct me if I am off base here.
>>> Regarding starting your next window with the previous one's final rst
>>> coordinates, that seems reasonable to me and I believe many people do
>> it
>>> that way, so you should be safe there. Are you using single atom
>> distance
>>> restraints, or center of mass distance restraints? CoM would probably
>> be
>>> preferable in your situation.
>>> Finally, regarding your force constant, I believe you have that
>> correct. I
>>> don't think any conversion is needed if you are calculating
>> distances,
>>> other than multiplying the force constant by two for WHAM.
>>> Hope this helps.
>>> Best regards,
>>> Zachary Fallon
>>>
>>> On Wed, Apr 17, 2019 at 6:59 AM Daniel Fernández Remacha
>>> <[3]dnlfr1994.gmail.com> wrote:
>>>
>>> Hi!Â
>>> Thank you very much for your answers!!
>>> It is our first time in the group using umbrella sampling and WHAM, so
>> any
>>> idea is absolutely appreciated.Â
>>> To give you a little more context of the system, our idea is to
>> separate a
>>> quite flexible peptide from the protein, from an initial distance of
>> about 7
>>> angstroms to 20 or so, to achieve a full separation of both molecules.Â
>>> For each increment of 0.2, r1 & r4 move 1unit above and below. We used
>> this
>>> because we thought this way the umbrella potential would be always
>>> symmetrical and equal for each window, allowing sampling with the same
>>> restrictions, and thus, pulling would not alter too much the structure
>> of
>>> the protein. It is more restrictive though.
>>> However, this needed what I mentioned at the end of my previous mail,
>> which
>>> makes sense to me but I am not sure if it is correct. Is it right to
>> start
>>> f.i. the second window (min-equil-prod) from the last prod.rst of the
>>> previous one? I suppose this may be unnecessary if y set r1= 1 and r4=
>> 90,
>>> since the borders always include de starting coordinates.
>>> I will try to simulate the umbrella with the conditions you mention and
>>> compare the results!
>>> Concerning WHAM, the only thing I suspect from is the restraint force.
>> The
>>> AMBER Advanced Tutorial 17 uses the same force as that used for the
>> umbrella
>>> and then multiplies by 2 and applies the conversion to degrees^2. Is
>> any
>>> conversion needed if distance is used instead of an angle? Am I right
>> if I
>>> say that using rk2=rk3=50 kcal/mol/ang, WHAM needs a force double of
>> what I
>>> used in AMBER? This is: 100kcal/mol/ang? No conversion needed?
>>> Thank you very much again
>>> Daniel Fernández
>>>
>>> El mar., 16 abr. 2019 21:23, Zachary Fallon
>>> <[4]zachary.fallon.stonybrook.edu> escribió:
>>>
>>> Hi Daniel,
>>> I only have a little experience with WHAM analysis, so I would wager
>> some of
>>> the more veteran users can weigh in here if I miss anything.
>>> Â Am I understanding you correctly that you varied your r1 and r4
>> values
>>> across the different windows? If you go back through the list serve
>> there is
>>> a good amount of discussion on what to set the r1 and r4 values, and
>>> I believe they should be constant across all of your windows ( I
>> faced
>>> similar issues as you when I varied my r1 and r4 across windows ). If
>> this
>>> is wrong I would love for someone to correct me. Â
>>> Additionally it is typically suggested that your r1 and r4 values be
>> 'far
>>> away' from your r2/r3 values, to ensure your potential is harmonic at
>> the
>>> sampled values of your defined coordinate. It is common to set r1 =
>> 1.0 or
>>> 0.0, and r4 to be some very large number (99.0 or 999). Check out the
>> Amber
>>> list serve archive for more details on how others have done this, like
>> this
>>> thread:Â [5]http://archive.ambermd.org/200705/0113.html.Â
>>> It is possible that having the r1 and r4 values vary across your
>> windows, as
>>> well as them being so close to your sampled region, is what is causing
>> you
>>> trouble. Without knowing much else about your system or setup, that is
>> where
>>> I would look first. Unfortunately that likely means rerunning your
>>> simulations.
>>> Finally, it is striking that your system requires a minimum of 50
>>> kcal/(mol*A^2) to maintain your fixed distances. That is a big energy
>>> penalty, and obviously it may make sense in the context of your
>> system, but
>>> it sticks out to me as unusual -- whatever barrier you are sampling
>> must be
>>> huge.Â
>>> Hope this helps, best of luck!
>>> --Zachary Fallon
>>>
>>> On Sat, Apr 13, 2019 at 9:20 AM Daniel Fernández Remacha
>>> <[6]dnlfr1994.gmail.com> wrote:
>>>
>>> Hi!!
>>> I am a student doing a Master Program in Pharmaceutical Chemistry
>> and I am
>>> writing to get some advice on understanding the results given by WHAM
>>> after
>>> doing an Umbrella Sampling of a dissociation process in a protein
>> (600
>>> residues approx.)-peptide interaction.
>>> To separate these molecules, a force constant of 50kcal/mol/ang2 (the
>>> smallest that can keep the distance reasonably steady) is set, and
>> r1 and
>>> r4 values fixed at -1 and +1 unit of each r2=r3 value for every (5 ns
>>> production) window. The increment of distance separation from window
>> to
>>> window is 0.2 angstroms.
>>> The combination of the MD gives a comprehensive separation movement
>> when
>>> visualized in VMD and umbrella sampling looks nice since the overlap
>> of
>>> the
>>> simulated windows looks adequate. It is much more full of dots than
>> the
>>> example of the tutorial.
>>> However, I am having difficulties to understand WHAM results.
>>> When looking at the result.dat (as it is called in the AMBER advanced
>>> tutorial 17) we get the following data.
>>> #Coor      Free       +/-     Â
>>>  Prob      +/-
>>> 7.492857Â Â 0.000000Â Â 0.000089Â Â 0.078953Â Â 0.000015
>>> 7.678571Â Â 0.021163Â Â 0.000114Â Â 0.076200Â Â 0.000018
>>> 7.864286Â Â 0.048638Â Â 0.000050Â Â 0.072768Â Â 0.000005
>>> 8.050000Â Â 0.035474Â Â 0.000201Â Â 0.074392Â Â 0.000012
>>> 8.235714Â Â 0.025529Â Â 0.000203Â Â 0.075644Â Â 0.000032
>>> 8.421429Â Â 0.074532Â Â 0.000235Â Â 0.069675Â Â 0.000034
>>> 8.607143Â Â 0.010492Â Â 0.000132Â Â 0.077576Â Â 0.000021
>>> 8.792857Â Â 0.031564Â Â 0.000090Â Â 0.074882Â Â 0.000009
>>> 8.978571Â Â 0.138947Â Â 0.000127Â Â 0.062539Â Â 0.000005
>>> 9.164286Â Â 0.160852Â Â 0.000055Â Â 0.060283Â Â 0.000007
>>> 9.350000Â Â 0.122887Â Â 0.000140Â Â 0.064246Â Â 0.000019
>>> 9.535714Â Â 0.063552Â Â 0.000138Â Â 0.070970Â Â 0.000021
>>> 9.721429Â Â 0.063538Â Â 0.000176Â Â 0.070971Â Â 0.000025
>>> 9.907143Â Â 0.064124Â Â 0.000422Â Â 0.070902Â Â 0.000016
>>> #Window    Free  +/-
>>> #0Â Â 0.000000Â Â 0.000000
>>> #1Â Â 0.000000Â Â 0.000000
>>> #2Â Â 0.000000Â Â 0.000000
>>> #3Â Â 0.015216Â Â 0.000000
>>> #4Â Â 0.000000Â Â 0.000000
>>> #5Â Â 0.000000Â Â 0.000000
>>> #6Â Â 0.000000Â Â 0.000000
>>> #7Â Â 0.000000Â Â 0.000000
>>> #8Â Â 0.010285Â Â 0.000000
>>> #9Â Â 0.000000Â Â 0.000000
>>> #10Â Â 0.000000Â Â 0.000000
>>> #11Â Â 0.000000Â Â 0.000000
>>> #12Â Â 0.000000Â Â 0.000000
>>> #13Â Â 0.034936Â Â 0.000000
>>> The main problem comes when looking at the Free energy, which is
>> used to
>>> see whether the system has converged or not. In the WHAM manual, it
>> is
>>> mentioned that when this F value is smaller than the specified
>> tolerance
>>> the system has converged. The tolerance value set for these results
>> is
>>> 10^-6.
>>> Moreover, when doing MonteCarlo Bootstrap analysis to look for
>>> similarities
>>> with the information given in the manual, we get 3 iterations; very
>>> different from the information given in the WHAM manual.
>>> I expected the system to be convergent due to a small increment
>> value,
>>> enough simulation time and good overlapping between windows.
>> However, I
>>> find strange to see these F values. The files present in the amber
>>> tutorial
>>> do not show these results ( #Window Free and +/-) and the WHAM
>> manual does
>>> not give information about the example used so we do not have a solid
>>> criterion to understand these results.
>>> I would be really grateful if you could help me understand the
>> meaning of
>>> these results for this system and provide any suggestion to perform a
>>> successful umbrella sampling.
>>> Thanks in advance,
>>> Daniel Fernández
>>> PD: We modified the Perl script for minimization, equilibration and
>>> production steps and our own script reads coordinates from the end
>> of the
>>> last production .rst file window performed; and not from the original
>>> coordinates as shown in the tutorial. I am not sure if this is
>> correct but
>>> we thought that this way the system would be less distorted when
>> reaching
>>> high distance values for separation.
>>> _______________________________________________
>>> AMBER mailing list
>>> [7]AMBER.ambermd.org
>>> [8]http://lists.ambermd.org/mailman/listinfo/amber
>>>
>>> --
>>>
>>> Zachary Fallon, PhD Candidate
>>> Dr. Carlos Simmerling Laboratory
>>> The Laufer Center for Physical and Quantitative Biology
>>> The Department of Chemistry, Stony Brook University
>>> Stony Brook, New York 11794
>>> Phone:Â [9](914) 703-1010Â Email:Â Â [10]
>> zachary.fallon.stonybrook.edu
>>>
>>> --
>>>
>>> Zachary Fallon, PhD Candidate
>>> Dr. Carlos Simmerling Laboratory
>>> The Laufer Center for Physical and Quantitative Biology
>>> The Department of Chemistry, Stony Brook University
>>> Stony Brook, New York 11794
>>> Phone:Â [11](914) 703-1010Â Email:Â Â [12]
>> zachary.fallon.stonybrook.edu
>>>
>>> --
>>>
>>> Zachary Fallon, PhD Candidate
>>> Dr. Carlos Simmerling Laboratory
>>> The Laufer Center for Physical and Quantitative Biology
>>> The Department of Chemistry, Stony Brook University
>>> Stony Brook, New York 11794
>>> Phone:Â [13](914) 703-1010Â Email:Â Â [14]
>> zachary.fallon.stonybrook.edu
>>>
>>> References
>>>
>>> 1. mailto:dnlfr1994.gmail.com
>>> 2. mailto:zachary.fallon.stonybrook.edu
>>> 3. mailto:dnlfr1994.gmail.com
>>> 4. mailto:zachary.fallon.stonybrook.edu
>>> 5. http://archive.ambermd.org/200705/0113.html
>>> 6. mailto:dnlfr1994.gmail.com
>>> 7. mailto:AMBER.ambermd.org
>>> 8. http://lists.ambermd.org/mailman/listinfo/amber
>>> 9. tel:(914)+703+1010
>>> 10. mailto:zachary.fallon.stonybrook.edu
>>> 11. tel:(914)+703+1010
>>> 12. mailto:zachary.fallon.stonybrook.edu
>>> 13. tel:(914)+703+1010
>>> 14. mailto:zachary.fallon.stonybrook.edu
>>>
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>>
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>>
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Received on Fri May 03 2019 - 08:30:03 PDT
