Re: [gmx-users] Can g_wham support using different temperature for different windows?
Hi, I think your PMF is asymetric because your peptide is asymetric and you don't sample enough. To get a symetric PMF, your peptide would have to sample all the possible conformations *and* orientations in each window. Thus it means that for the windows in the center of the bilayer (where you say it's extended and interacts with the two monolayers) it'll have to rotate completely the other way round. This event will probably be *very* rare because you have to translocate positive charges across the membrane which cost ~ 40 to 50 kJ/mol (see the PMF of Lys+ and Arg+ in 10.1021/ct700324x). So as suggested by Chris, Justin and Xavier, you'll have to sample way more than 100 ns per window. I think you should go at least to the microsecond time scale (or more?). Or maybe starting from different initial conformations/orientations for a given window and then concatenate the different trajectories? Also consider the remark of Xavier, TM or interfacial peptides are most of the time alpha-helical within the membrane. So far in literature, PMFs of a whole peptide across a bilayer were done by restraining the peptide in a helical conformation (e.g. 10.1016/j.bpj.2010.12.3682). It is anyway a very difficult problem (and probably impossible at atomistic resolution) to get a converged PMF for a whole peptide (e.g. 10.1016/j.bpj.2009.03.059). Ciao, Patrick Le 23/02/2011 05:25, Jianguo Li a écrit : Sorry, why do you think the PMF should be asymmetric? I pulled my peptide from d=9nm (above the membrane) to d=-3nm (below the membrane) and I did windowed umbrella sampling in the range of d=-1.05nm to d=9nm. At least the PMF should be symmetric with respect of the bilayer center in the range of d=[-1.05nm 1.05nm], something like a guassian distribution. But I got asymmetric PMF in this region. I also did reverse pulling starting from the peptide below the membrane ending with the peptide above the membrane. And the subsequent PMF of reversed pulling is also asymmetic. I have position restrains of the phosphate beads of the lipids in z-direction. So the membrane should be stable in REMD. But as you mentioned, if peptide is truly stuck in this orientation, REMD may be not useful. I will do a single simulation first at a higher temperature (e.g., 400K) in those bad windows to see if the peptide conformations are fully sampled. Cheers, Jianguo *From:* Justin A. Lemkul jalem...@vt.edu *To:* Gromacs Users' List gmx-users@gromacs.org *Sent:* Wednesday, 23 February 2011 10:24:46 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thank you, Justin. Actually I did windowed umbrella simulations from d=-1.05nm to d=9nm. Since I think there is no problem in the region out of the membrane, so I only show the configurations within the membrane. My objective is to access the free energy barrier of the peptide translocate the negatively charged membrane. The problem is that the PMF is not symmetric with respect to the bilayer center due to the unconverged simulations. I would argue that the PMF is not symmetric because your reaction coordinate is not symmetric. How can you calculate a free energy of crossing a charged membrane when your peptide does not cross the membrane? What I proposed earlier was to obtain configurations at equal distances above and below the membrane (arbitrary in a periodic system, but hopefully you get the idea). If you can extract the peptide to the point where it is liberated from the membrane in the negative direction, I'd suspect you could solve your problem. Since g_wham does not support different temperatures in different windows, to increase the converges, I will probably consider to do REMD in those bad windows. This technique might work, provided you don't destabilize the membrane, but if the peptide is truly stuck in this orientation, I doubt that limited-range REMD would be very useful. -Justin Cheers Jianguo *From:* Justin A. Lemkul jalem...@vt.edu mailto:jalem...@vt.edu *To:* Gromacs Users' List gmx-users@gromacs.org mailto:gmx-users@gromacs.org *Sent:* Tuesday, 22 February 2011 21:10:08 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thanks Justin and Chris and sorry for confusing interpretation. Let me make it more clear. My peptide is flexible Martini beads, and highly positively charged. My membrane is a mixture of negatively charged lipids (25%) and zitterionic lipids(75%). So there is strong electrostatic attraction between peptide and membrane. To get the PMF, I did the following: (1) I did pulling simulation along (0 0 -1) direction to pull my peptide across the membrane. Then I got different configurations corresponding to different windows
Re: [gmx-users] Can g_wham support using different temperature for different windows?
On Feb 23, 2011, at 3:21 AM, Jianguo Li wrote: Thank you for the the useful information, XAvier. My peptide is highly positively charged, 18 AA with +12 charges. Other of my group members told me their NMR experiment in water indicates the peptide conformation is very dynamics. Actually I also did peptide refolding using REMD in water, and I found it is flexible and has no stable structure in water, except some instantaneously helical structures. In addition, my peptide consists of two branches connected by unnatural peptide bond, so the backbone is discontinuous, and also because of the high charges, I assume the peptide doesn't form helcial structure in the negatively charged membrane. Therefore I didn't put any constraints in the peptide to keep the secondary structure of the peptide. I know there are assumptions in my model, but I have no other information to increase the accuracy of the model. In fact, when I am doing REMD folding simulations using Gromos53a6 and CHARMM27 with cMap, I got different results. But the common thing is that both results seems to indicate the peptide is filexbile in water without stable secondary structure. Then I used MARTINI FF with flexible structure, just to find some general features. I will try your suggestion, doing REMD in those bad windows. And the reference you mentioned is very useful, I will take a look at them :-) Another question: Suppose some other tools support using different temperatures in different windows, as you mentioned, if 500K is too high to have a significant contribution to the probability of 300K, can I do a series of simulation in a certain window with different temparatures (e.g. 300K, 350K, 400K,450K, 500K). In such cases, in each window, I need to do 5 simulations, which will be much cheaper than doing REMD in that window. It would be computationally cheaper but this is assuming that you'd get the info you are looking for within these simulations and again the weight of the conformations from 400/450/500 K at 300 K is questionable. Note also that the conformations sampled at high temperature with position restrains on the lipids to avoid deformation will be difficult to interpret! Cheers Jianguo From: XAvier Periole x.peri...@rug.nl To: Discussion list for GROMACS users gmx-users@gromacs.org Sent: Tuesday, 22 February 2011 21:18:12 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? A few notes: - the original method (Kumar-JCC-1992) that inspired wham was actually developed to mix different temperature simulations. It is however not clear for the type of system you are simulating how much a 500K simulation would be useful to improve the sampling at 300 K or so. The reason is that the enthalpy difference between the two systems is so high that the probability that a conformation from a 500K simulation would contribute to sampling at 300K is really low. It would much more efficient for systems with implicit solvent for which the energy of the system does not vary so much with the temperature. One could look at Chodera-JCTC-2007 and ref therein for a few examples. - I would think that a REMD simulation would be more useful. No need to run 30 replicas to very hight temperature! A bilayer at 500K might get funny. - Martini force field for flexible regions of protein should not be trusted ... or really interpreted with a lot of reserve. The coil definition is simply something flexible with absolutely no guaranty that it could be representing some thing even close to reality, which we have only an approximate idea of what it is! - A peptide in a bilayer has a very high chance to get into a helical conformation. Do you think it is reasonable to keep it flexible? - As noted by Justin and Chris, you definitely have a problem of convergence ... I am not sure how many converged examples of PMFs of peptide crossing a bilayer are out in the literature (Justin?) but from our experience with Martini it does take an awful lot of time to really get convergence. For you system I would expect at least a microsecond for the windows where sampling is an issue. As an example, we saw significant differences on a PMF between two simple helices up to 8 us ... and no charges were involved. This might be a lot pessimistic but you should not get fooled by a CG model. Martini is really good for a lot of things but other things should really but be looked at carefully. XAvier. On Feb 22, 2011, at 9:12 AM, Jianguo Li wrote: Sorry I forgot to attach my mdp files. Here is the mdp file for pulling simulaition: - -- gmx-users mailing listgmx-users@gromacs.org http://lists.gromacs.org/mailman/listinfo/gmx-users Please search the archive at http://www.gromacs.org/Support/Mailing_Lists/Search before posting! Please don't
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Thanks for your comments, Patric. You are right. The energy barrier is too high for charged groups to translocate the hydrophobic region of the membrane. And my peptide contains 12 positively charge residues (ARG and LYS), therefore it is unlikely to sample those translocation. I am considering to extend my simulations to microsecond level or longer or use REMD. Cheers, Jianguo From: Patrick Fuchs patrick.fu...@univ-paris-diderot.fr To: Discussion list for GROMACS users gmx-users@gromacs.org Sent: Wednesday, 23 February 2011 19:04:05 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? Hi, I think your PMF is asymetric because your peptide is asymetric and you don't sample enough. To get a symetric PMF, your peptide would have to sample all the possible conformations *and* orientations in each window. Thus it means that for the windows in the center of the bilayer (where you say it's extended and interacts with the two monolayers) it'll have to rotate completely the other way round. This event will probably be *very* rare because you have to translocate positive charges across the membrane which cost ~ 40 to 50 kJ/mol (see the PMF of Lys+ and Arg+ in 10.1021/ct700324x). So as suggested by Chris, Justin and Xavier, you'll have to sample way more than 100 ns per window. I think you should go at least to the microsecond time scale (or more?). Or maybe starting from different initial conformations/orientations for a given window and then concatenate the different trajectories? Also consider the remark of Xavier, TM or interfacial peptides are most of the time alpha-helical within the membrane. So far in literature, PMFs of a whole peptide across a bilayer were done by restraining the peptide in a helical conformation (e.g. 10.1016/j.bpj.2010.12.3682). It is anyway a very difficult problem (and probably impossible at atomistic resolution) to get a converged PMF for a whole peptide (e.g. 10.1016/j.bpj.2009.03.059). Ciao, Patrick Le 23/02/2011 05:25, Jianguo Li a écrit : Sorry, why do you think the PMF should be asymmetric? I pulled my peptide from d=9nm (above the membrane) to d=-3nm (below the membrane) and I did windowed umbrella sampling in the range of d=-1.05nm to d=9nm. At least the PMF should be symmetric with respect of the bilayer center in the range of d=[-1.05nm 1.05nm], something like a guassian distribution. But I got asymmetric PMF in this region. I also did reverse pulling starting from the peptide below the membrane ending with the peptide above the membrane. And the subsequent PMF of reversed pulling is also asymmetic. I have position restrains of the phosphate beads of the lipids in z-direction. So the membrane should be stable in REMD. But as you mentioned, if peptide is truly stuck in this orientation, REMD may be not useful. I will do a single simulation first at a higher temperature (e.g., 400K) in those bad windows to see if the peptide conformations are fully sampled. Cheers, Jianguo *From:* Justin A. Lemkul jalem...@vt.edu *To:* Gromacs Users' List gmx-users@gromacs.org *Sent:* Wednesday, 23 February 2011 10:24:46 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thank you, Justin. Actually I did windowed umbrella simulations from d=-1.05nm to d=9nm. Since I think there is no problem in the region out of the membrane, so I only show the configurations within the membrane. My objective is to access the free energy barrier of the peptide translocate the negatively charged membrane. The problem is that the PMF is not symmetric with respect to the bilayer center due to the unconverged simulations. I would argue that the PMF is not symmetric because your reaction coordinate is not symmetric. How can you calculate a free energy of crossing a charged membrane when your peptide does not cross the membrane? What I proposed earlier was to obtain configurations at equal distances above and below the membrane (arbitrary in a periodic system, but hopefully you get the idea). If you can extract the peptide to the point where it is liberated from the membrane in the negative direction, I'd suspect you could solve your problem. Since g_wham does not support different temperatures in different windows, to increase the converges, I will probably consider to do REMD in those bad windows. This technique might work, provided you don't destabilize the membrane, but if the peptide is truly stuck in this orientation, I doubt that limited-range REMD would be very useful. -- gmx-users mailing listgmx-users@gromacs.org http://lists.gromacs.org/mailman/listinfo/gmx-users Please search the archive at http://www.gromacs.org/Support/Mailing_Lists/Search before posting! Please don't post (un
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Dear all, Thank you all for your suggestions or comments to my problem. Now I am planning to extend my simulations or using REMD in those bad windows to get converged PMF. I have another question: if I extend the umbrellar simulation to 1 microsecond only in those problematic windows, while running shorter simulation (e.g., 100 ns) in those windows far away from the membrane. Does g_wham accpet using diffrent simulation time for different windows? Thank you in again, Cheers, Jianguo From: XAvier Periole x.peri...@rug.nl To: Discussion list for GROMACS users gmx-users@gromacs.org Sent: Wednesday, 23 February 2011 20:59:18 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? On Feb 23, 2011, at 3:21 AM, Jianguo Li wrote: Thank you for the the useful information, XAvier. My peptide is highly positively charged, 18 AA with +12 charges. Other of my group members told me their NMR experiment in water indicates the peptide conformation is very dynamics. Actually I also did peptide refolding using REMD in water, and I found it is flexible and has no stable structure in water, except some instantaneously helical structures. In addition, my peptide consists of two branches connected by unnatural peptide bond, so the backbone is discontinuous, and also because of the high charges, I assume the peptide doesn't form helcial structure in the negatively charged membrane. Therefore I didn't put any constraints in the peptide to keep the secondary structure of the peptide. I know there are assumptions in my model, but I have no other information to increase the accuracy of the model. In fact, when I am doing REMD folding simulations using Gromos53a6 and CHARMM27 with cMap, I got different results. But the common thing is that both results seems to indicate the peptide is filexbile in water without stable secondary structure. Then I used MARTINI FF with flexible structure, just to find some general features. I will try your suggestion, doing REMD in those bad windows. And the reference you mentioned is very useful, I will take a look at them :-) Another question: Suppose some other tools support using different temperatures in different windows, as you mentioned, if 500K is too high to have a significant contribution to the probability of 300K, can I do a series of simulation in a certain window with different temparatures (e.g. 300K, 350K, 400K,450K, 500K). In such cases, in each window, I need to do 5 simulations, which will be much cheaper than doing REMD in that window. It would be computationally cheaper but this is assuming that you'd get the info you are looking for within these simulations and again the weight of the conformations from 400/450/500 K at 300 K is questionable. Note also that the conformations sampled at high temperature with position restrains on the lipids to avoid deformation will be difficult to interpret! Cheers Jianguo From: XAvier Periole x.peri...@rug.nl To: Discussion list for GROMACS users gmx-users@gromacs.org Sent: Tuesday, 22 February 2011 21:18:12 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? A few notes: - the original method (Kumar-JCC-1992) that inspired wham was actually developed to mix different temperature simulations. It is however not clear for the type of system you are simulating how much a 500K simulation would be useful to improve the sampling at 300 K or so. The reason is that the enthalpy difference between the two systems is so high that the probability that a conformation from a 500K simulation would contribute to sampling at 300K is really low. It would much more efficient for systems with implicit solvent for which the energy of the system does not vary so much with the temperature. One could look at Chodera-JCTC-2007 and ref therein for a few examples. - I would think that a REMD simulation would be more useful. No need to run 30 replicas to very hight temperature! A bilayer at 500K might get funny. - Martini force field for flexible regions of protein should not be trusted ... or really interpreted with a lot of reserve. The coil definition is simply something flexible with absolutely no guaranty that it could be representing some thing even close to reality, which we have only an approximate idea of what it is! - A peptide in a bilayer has a very high chance to get into a helical conformation. Do you think it is reasonable to keep it flexible? - As noted by Justin and Chris, you definitely have a problem of convergence ... I am not sure how many converged examples of PMFs of peptide crossing a bilayer are out in the literature (Justin?) but from our experience with Martini it does take an awful lot of time to really get convergence. For you system I would expect at least a microsecond for the windows where sampling
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Jianguo Li wrote: Dear all, Thank you all for your suggestions or comments to my problem. Now I am planning to extend my simulations or using REMD in those bad windows to get converged PMF. I have another question: if I extend the umbrellar simulation to 1 microsecond only in those problematic windows, while running shorter simulation (e.g., 100 ns) in those windows far away from the membrane. Does g_wham accpet using diffrent simulation time for different windows? Theoretically, but if you are going to discard any of the initial time (that is, equilibration in each window) by using the -b option, you'll have to make sure that you don't eliminate windows by doing so, i.e. 100 ns in some windows and 1 us in others, using g_wham -b 20 would completely neglect any windows with shorter time and basically make the PMF curve useless. -Justin Thank you in again, Cheers, Jianguo *From:* XAvier Periole x.peri...@rug.nl *To:* Discussion list for GROMACS users gmx-users@gromacs.org *Sent:* Wednesday, 23 February 2011 20:59:18 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? On Feb 23, 2011, at 3:21 AM, Jianguo Li wrote: Thank you for the the useful information, XAvier. My peptide is highly positively charged, 18 AA with +12 charges. Other of my group members told me their NMR experiment in water indicates the peptide conformation is very dynamics. Actually I also did peptide refolding using REMD in water, and I found it is flexible and has no stable structure in water, except some instantaneously helical structures. In addition, my peptide consists of two branches connected by unnatural peptide bond, so the backbone is discontinuous, and also because of the high charges, I assume the peptide doesn't form helcial structure in the negatively charged membrane. Therefore I didn't put any constraints in the peptide to keep the secondary structure of the peptide. I know there are assumptions in my model, but I have no other information to increase the accuracy of the model. In fact, when I am doing REMD folding simulations using Gromos53a6 and CHARMM27 with cMap, I got different results. But the common thing is that both results seems to indicate the peptide is filexbile in water without stable secondary structure. Then I used MARTINI FF with flexible structure, just to find some general features. I will try your suggestion, doing REMD in those bad windows. And the reference you mentioned is very useful, I will take a look at them :-) Another question: Suppose some other tools support using different temperatures in different windows, as you mentioned, if 500K is too high to have a significant contribution to the probability of 300K, can I do a series of simulation in a certain window with different temparatures (e.g. 300K, 350K, 400K,450K, 500K). In such cases, in each window, I need to do 5 simulations, which will be much cheaper than doing REMD in that window. It would be computationally cheaper but this is assuming that you'd get the info you are looking for within these simulations and again the weight of the conformations from 400/450/500 K at 300 K is questionable. Note also that the conformations sampled at high temperature with position restrains on the lipids to avoid deformation will be difficult to interpret! Cheers Jianguo *From:* XAvier Periole x.peri...@rug.nl mailto:x.peri...@rug.nl *To:* Discussion list for GROMACS users gmx-users@gromacs.org mailto:gmx-users@gromacs.org *Sent:* Tuesday, 22 February 2011 21:18:12 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? A few notes: - the original method (Kumar-JCC-1992) that inspired wham was actually developed to mix different temperature simulations. It is however not clear for the type of system you are simulating how much a 500K simulation would be useful to improve the sampling at 300 K or so. The reason is that the enthalpy difference between the two systems is so high that the probability that a conformation from a 500K simulation would contribute to sampling at 300K is really low. It would much more efficient for systems with implicit solvent for which the energy of the system does not vary so much with the temperature. One could look at Chodera-JCTC-2007 and ref therein for a few examples. - I would think that a REMD simulation would be more useful. No need to run 30 replicas to very hight temperature! A bilayer at 500K might get funny. - Martini force field for flexible regions of protein should not be trusted ... or really interpreted with a lot of reserve. The coil definition is simply something flexible with absolutely no guaranty that it could be representing some thing even close to reality, which we have
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Sorry I forgot to attach my mdp files. Here is the mdp file for pulling simulaition: - title= Martini cpp = /usr/bin/cpp define = -DPOSRES_LIP integrator = md ; start time and timestep in ps tinit= 0.0 dt = 0.02 nsteps = 500 ; number of steps for center of mass motion removal = nstcomm = 10 comm-grps= nstxout = 5000 nstvout = 50 nstfout = 0 ; Output frequency for energies to log file and energy file = nstlog = 1000 nstenergy= 1000 ; Output frequency and precision for xtc file = nstxtcout= 1000 xtc_precision= 100 nstlist = 10 ; ns algorithm (simple or grid) = ns_type = grid ; Periodic boundary conditions: xyz or none = pbc = xyz ; nblist cut-off = rlist= 1.4 coulombtype = PME rcoulomb = 1.4 fourierspacing = 0.12 pme_order = 4 ewald_rtol = 1e-5 optimize_fft = yes ; Dielectric constant (DC) for cut-off or DC of reaction field = epsilon_r= 15 ; Method for doing Van der Waals = vdw_type = Shift ; cut-off lengths= rvdw_switch = 0.9 rvdw = 1.2 ; Apply long range dispersion corrections for Energy and Pressure = DispCorr = No ; Temperature coupling = tcoupl = V-rescale ; Groups to couple separately = tc-grps = Protein_lipid Sol_Ion ; Time constant (ps) and reference temperature (K) = tau_t= 1.5 1.5 ref_t= 310 310 ; Pressure coupling = Pcoupl = Parrinello-Rahman Pcoupltype = semiisotropic ; Time constant (ps), compressibility (1/bar) and reference P (bar) = tau_p= 10.0 10.0 compressibility = 3e-5 3e-5 ref_p= 1.0 1.0 constraints = none ; Type of constraint algorithm = constraint_algorithm = Lincs ; Do not constrain the start configuration = unconstrained_start = no ; Highest order in the expansion of the constraint coupling matrix = lincs_order = 4 ; Lincs will write a warning to the stderr if in one step a bond = ; rotates over more degrees than = lincs_warnangle = 90 ; pull staff ; pull staff pull= umbrella pull_geometry = position pull_vec1= 0 0 -1 pull_dim = N N Y pull_start = no ; define initial COM distance 0 pull_ngroups= 1 pull_group0 = lipid1 pull_group1 = Protein pull_init1 = 0.0 0.0 4.50 pull_rate1 = 0.001 ; 0.01 nm per ps = 10 nm per ns pull_k1 = 1000 ; kJ mol^-1 nm^-2 Here is the pull part of the mpd file for the windowed umbrella sampling simulation, other part of the mdp file are same as that of pulling simulation. pull= umbrella pull_geometry = position pull_vec1= 0 0 -1 pull_dim = N N Y pull_start = no ; define initial COM distance 0 pull_ngroups= 1 pull_group0 = lipid1 pull_group1 = Protein pull_init1 = 0.0 0.0 1.2 pull_k1 = 1000 ; kJ mol^-1 nm^-2 Cheers, Jianguo From: Jianguo Li ljg...@yahoo.com.sg To: jalem...@vt.edu; Discussion list for GROMACS users gmx-users@gromacs.org Sent: Tuesday, 22 February 2011 14:27:34 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? Thanks Justin and Chris and sorry for confusing interpretation. Let me make it more clear. My peptide is flexible Martini beads, and highly positively charged. My membrane is a mixture of negatively charged lipids (25%) and zitterionic lipids(75%). So there is strong electrostatic attraction between peptide and membrane. To get the PMF, I did the following: (1) I did pulling simulation along (0 0 -1) direction to pull my peptide across the membrane. Then I got different configurations corresponding to different windows along the reaction coordinates, which is the z-distance between peptide and membrane. This figure (http://www.flickr.com/photos/lijg/5467080971/) shows some of the configurations at certain reaction coordinates. (2) In each window, I used the corresponding configuration that generated by the pulling simulation as initial input and run umbrella sampling. The size of each window is 0.15 nm, but close to the bilayer cneter (e.g., -0.6d0.6), I have increased number of windows so that the width of the window is to be 0.05 or 0.1 nm, I also tried to use different force
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Jianguo Li wrote: Thanks Justin and Chris and sorry for confusing interpretation. Let me make it more clear. My peptide is flexible Martini beads, and highly positively charged. My membrane is a mixture of negatively charged lipids (25%) and zitterionic lipids(75%). So there is strong electrostatic attraction between peptide and membrane. To get the PMF, I did the following: (1) I did pulling simulation along (0 0 -1) direction to pull my peptide across the membrane. Then I got different configurations corresponding to different windows along the reaction coordinates, which is the z-distance between peptide and membrane. This figure (http://www.flickr.com/photos/lijg/5467080971/) shows some of the configurations at certain reaction coordinates. Are you not sampling configurations outside of the membrane (i.e. in water)? I would think that would solve your problem. You don't show any configurations in which the peptide is completely dissociated from the membrane. I don't know your objectives, but I would think that if you could completely extract the peptide from the membrane after passing through it, this would solve your problem. (2) In each window, I used the corresponding configuration that generated by the pulling simulation as initial input and run umbrella sampling. The size of each window is 0.15 nm, but close to the bilayer cneter (e.g., -0.6d0.6), I have increased number of windows so that the width of the window is to be 0.05 or 0.1 nm, I also tried to use different force constant in these windows. From the figure (http://www.flickr.com/photos/lijg/5467080971/) , we can classify the peptide conformation to be either extended (interacting with two bilayers) or compact (interacting with only one bilayer). Ideally, the peptide conformation should be similar for d=x and d=-x. The problem is that the configuration of peptide is not symmetric with respect to the bilayer center. For example, the peptide configuration is compact at d=0.6 and d=0.9, but the peptide is extended at d=-0.6 and d=-0.9. This leads Hysteresis. If I use g_wham to generate PMF, then the PMF is not symmetric with respect to the bilayer center. Using more number of windows and different force constant did not remove the problem. In my opinion, at least in some windows, the peptide should sample both compact and extended structure. But what I found is that the windowed Don't pre-judge the model :) Also, as I said before, there is no reason to suspect that MARTINI will produce any meaningful secondary structure changes. It was not parameterized to do so. umbrella simulation depends on the initial peptide conformation. If the initial peptide conformation is compact, then after 100 ns, it is still compact; if the initial peptide in that window is extended, the final configuration is also extended. I also tried to run longer equilibrium time (e.g., 200 ns), but the problem still exists. Sounds like a limitation of the force field model. My question is how to increase sampling of the peptide conformation? I just think of two choices: (1) use high temperature (e.g., 500K) at those bad windows. As I mentioned, I am wondering if g_wham can unbias the effect of using different temperatures in different windows. (2) use REMD in those bad windows. These need a lot of computational resources. Neither of these will be useful in generating a sensible PMF curve. WHAM needs a single temperature for proper weighting. If you start including different temperatures in different regions of phase space, I would imagine the weighting would be completely incorrect. Note that SMD is not the only option for generating starting configurations. If you think that certain orientations or configurations are correct, you can build them yourself, but keep in mind that you'll have to justify this procedure to a skeptical audience. -Justin Is there any other method to deal with the insufficient sampling? Any suggestions are welcome, thanks for your time reading this email! Cheers, Jianguo *From:* Justin A. Lemkul jalem...@vt.edu *To:* Gromacs Users' List gmx-users@gromacs.org *Sent:* Tuesday, 22 February 2011 11:13:05 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thanks for your comments, Justin. Using timestep of 20 fs, in each window the simulation runs for 100 ns CG time. The pulling rate is 0.001 nm/ps. Is it too fast? Let me clarify things, since I'm not convinced I understand your procedure. You generate a series of configurations with 0.001 nm/ps pulling, but then how many windows do you generate for independent simulations? What are your .mdp parameters during those windows? The pull rate should be 0 during the actual umbrella sampling, to restrain the peptide within the window. What force constant(s) do
Re: [gmx-users] Can g_wham support using different temperature for different windows?
A few notes: - the original method (Kumar-JCC-1992) that inspired wham was actually developed to mix different temperature simulations. It is however not clear for the type of system you are simulating how much a 500K simulation would be useful to improve the sampling at 300 K or so. The reason is that the enthalpy difference between the two systems is so high that the probability that a conformation from a 500K simulation would contribute to sampling at 300K is really low. It would much more efficient for systems with implicit solvent for which the energy of the system does not vary so much with the temperature. One could look at Chodera-JCTC-2007 and ref therein for a few examples. - I would think that a REMD simulation would be more useful. No need to run 30 replicas to very hight temperature! A bilayer at 500K might get funny. - Martini force field for flexible regions of protein should not be trusted ... or really interpreted with a lot of reserve. The coil definition is simply something flexible with absolutely no guaranty that it could be representing some thing even close to reality, which we have only an approximate idea of what it is! - A peptide in a bilayer has a very high chance to get into a helical conformation. Do you think it is reasonable to keep it flexible? - As noted by Justin and Chris, you definitely have a problem of convergence ... I am not sure how many converged examples of PMFs of peptide crossing a bilayer are out in the literature (Justin?) but from our experience with Martini it does take an awful lot of time to really get convergence. For you system I would expect at least a microsecond for the windows where sampling is an issue. As an example, we saw significant differences on a PMF between two simple helices up to 8 us ... and no charges were involved. This might be a lot pessimistic but you should not get fooled by a CG model. Martini is really good for a lot of things but other things should really but be looked at carefully. XAvier. On Feb 22, 2011, at 9:12 AM, Jianguo Li wrote: Sorry I forgot to attach my mdp files. Here is the mdp file for pulling simulaition: - -- gmx-users mailing listgmx-users@gromacs.org http://lists.gromacs.org/mailman/listinfo/gmx-users Please search the archive at http://www.gromacs.org/Support/Mailing_Lists/Search before posting! Please don't post (un)subscribe requests to the list. Use the www interface or send it to gmx-users-requ...@gromacs.org. Can't post? Read http://www.gromacs.org/Support/Mailing_Lists
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Thank you, Justin. Actually I did windowed umbrella simulations from d=-1.05nm to d=9nm. Since I think there is no problem in the region out of the membrane, so I only show the configurations within the membrane. My objective is to access the free energy barrier of the peptide translocate the negatively charged membrane. The problem is that the PMF is not symmetric with respect to the bilayer center due to the unconverged simulations. Since g_wham does not support different temperatures in different windows, to increase the converges, I will probably consider to do REMD in those bad windows. Cheers Jianguo From: Justin A. Lemkul jalem...@vt.edu To: Gromacs Users' List gmx-users@gromacs.org Sent: Tuesday, 22 February 2011 21:10:08 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thanks Justin and Chris and sorry for confusing interpretation. Let me make it more clear. My peptide is flexible Martini beads, and highly positively charged. My membrane is a mixture of negatively charged lipids (25%) and zitterionic lipids(75%). So there is strong electrostatic attraction between peptide and membrane. To get the PMF, I did the following: (1) I did pulling simulation along (0 0 -1) direction to pull my peptide across the membrane. Then I got different configurations corresponding to different windows along the reaction coordinates, which is the z-distance between peptide and membrane. This figure (http://www.flickr.com/photos/lijg/5467080971/) shows some of the configurations at certain reaction coordinates. Are you not sampling configurations outside of the membrane (i.e. in water)? I would think that would solve your problem. You don't show any configurations in which the peptide is completely dissociated from the membrane. I don't know your objectives, but I would think that if you could completely extract the peptide from the membrane after passing through it, this would solve your problem. (2) In each window, I used the corresponding configuration that generated by the pulling simulation as initial input and run umbrella sampling. The size of each window is 0.15 nm, but close to the bilayer cneter (e.g., -0.6d0.6), I have increased number of windows so that the width of the window is to be 0.05 or 0.1 nm, I also tried to use different force constant in these windows. From the figure (http://www.flickr.com/photos/lijg/5467080971/) , we can classify the peptide conformation to be either extended (interacting with two bilayers) or compact (interacting with only one bilayer). Ideally, the peptide conformation should be similar for d=x and d=-x. The problem is that the configuration of peptide is not symmetric with respect to the bilayer center. For example, the peptide configuration is compact at d=0.6 and d=0.9, but the peptide is extended at d=-0.6 and d=-0.9. This leads Hysteresis. If I use g_wham to generate PMF, then the PMF is not symmetric with respect to the bilayer center. Using more number of windows and different force constant did not remove the problem. In my opinion, at least in some windows, the peptide should sample both compact and extended structure. But what I found is that the windowed Don't pre-judge the model :) Also, as I said before, there is no reason to suspect that MARTINI will produce any meaningful secondary structure changes. It was not parameterized to do so. umbrella simulation depends on the initial peptide conformation. If the initial peptide conformation is compact, then after 100 ns, it is still compact; if the initial peptide in that window is extended, the final configuration is also extended. I also tried to run longer equilibrium time (e.g., 200 ns), but the problem still exists. Sounds like a limitation of the force field model. My question is how to increase sampling of the peptide conformation? I just think of two choices: (1) use high temperature (e.g., 500K) at those bad windows. As I mentioned, I am wondering if g_wham can unbias the effect of using different temperatures in different windows. (2) use REMD in those bad windows. These need a lot of computational resources. Neither of these will be useful in generating a sensible PMF curve. WHAM needs a single temperature for proper weighting. If you start including different temperatures in different regions of phase space, I would imagine the weighting would be completely incorrect. Note that SMD is not the only option for generating starting configurations. If you think that certain orientations or configurations are correct, you can build them yourself, but keep in mind that you'll have to justify this procedure to a skeptical audience. -Justin Is there any other method to deal with the insufficient sampling? Any suggestions are welcome, thanks for your time reading this email! Cheers, Jianguo
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Thank you for the the useful information, XAvier. My peptide is highly positively charged, 18 AA with +12 charges. Other of my group members told me their NMR experiment in water indicates the peptide conformation is very dynamics. Actually I also did peptide refolding using REMD in water, and I found it is flexible and has no stable structure in water, except some instantaneously helical structures. In addition, my peptide consists of two branches connected by unnatural peptide bond, so the backbone is discontinuous, and also because of the high charges, I assume the peptide doesn't form helcial structure in the negatively charged membrane. Therefore I didn't put any constraints in the peptide to keep the secondary structure of the peptide. I know there are assumptions in my model, but I have no other information to increase the accuracy of the model. In fact, when I am doing REMD folding simulations using Gromos53a6 and CHARMM27 with cMap, I got different results. But the common thing is that both results seems to indicate the peptide is filexbile in water without stable secondary structure. Then I used MARTINI FF with flexible structure, just to find some general features. I will try your suggestion, doing REMD in those bad windows. And the reference you mentioned is very useful, I will take a look at them :-) Another question: Suppose some other tools support using different temperatures in different windows, as you mentioned, if 500K is too high to have a significant contribution to the probability of 300K, can I do a series of simulation in a certain window with different temparatures (e.g. 300K, 350K, 400K,450K, 500K). In such cases, in each window, I need to do 5 simulations, which will be much cheaper than doing REMD in that window. Cheers Jianguo From: XAvier Periole x.peri...@rug.nl To: Discussion list for GROMACS users gmx-users@gromacs.org Sent: Tuesday, 22 February 2011 21:18:12 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? A few notes: - the original method (Kumar-JCC-1992) that inspired wham was actually developed to mix different temperature simulations. It is however not clear for the type of system you are simulating how much a 500K simulation would be useful to improve the sampling at 300 K or so. The reason is that the enthalpy difference between the two systems is so high that the probability that a conformation from a 500K simulation would contribute to sampling at 300K is really low. It would much more efficient for systems with implicit solvent for which the energy of the system does not vary so much with the temperature. One could look at Chodera-JCTC-2007 and ref therein for a few examples. - I would think that a REMD simulation would be more useful. No need to run 30 replicas to very hight temperature! A bilayer at 500K might get funny. - Martini force field for flexible regions of protein should not be trusted ... or really interpreted with a lot of reserve. The coil definition is simply something flexible with absolutely no guaranty that it could be representing some thing even close to reality, which we have only an approximate idea of what it is! - A peptide in a bilayer has a very high chance to get into a helical conformation. Do you think it is reasonable to keep it flexible? - As noted by Justin and Chris, you definitely have a problem of convergence ... I am not sure how many converged examples of PMFs of peptide crossing a bilayer are out in the literature (Justin?) but from our experience with Martini it does take an awful lot of time to really get convergence. For you system I would expect at least a microsecond for the windows where sampling is an issue. As an example, we saw significant differences on a PMF between two simple helices up to 8 us ... and no charges were involved. This might be a lot pessimistic but you should not get fooled by a CG model. Martini is really good for a lot of things but other things should really but be looked at carefully. XAvier. On Feb 22, 2011, at 9:12 AM, Jianguo Li wrote: Sorry I forgot to attach my mdp files. Here is the mdp file for pulling simulaition: - -- gmx-users mailing listgmx-users@gromacs.org http://lists.gromacs.org/mailman/listinfo/gmx-users Please search the archive at http://www.gromacs.org/Support/Mailing_Lists/Search before posting! Please don't post (un)subscribe requests to the list. Use the www interface or send it to gmx-users-requ...@gromacs.org. Can't post? Read http://www.gromacs.org/Support/Mailing_Lists
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Jianguo Li wrote: Thank you, Justin. Actually I did windowed umbrella simulations from d=-1.05nm to d=9nm. Since I think there is no problem in the region out of the membrane, so I only show the configurations within the membrane. My objective is to access the free energy barrier of the peptide translocate the negatively charged membrane. The problem is that the PMF is not symmetric with respect to the bilayer center due to the unconverged simulations. I would argue that the PMF is not symmetric because your reaction coordinate is not symmetric. How can you calculate a free energy of crossing a charged membrane when your peptide does not cross the membrane? What I proposed earlier was to obtain configurations at equal distances above and below the membrane (arbitrary in a periodic system, but hopefully you get the idea). If you can extract the peptide to the point where it is liberated from the membrane in the negative direction, I'd suspect you could solve your problem. Since g_wham does not support different temperatures in different windows, to increase the converges, I will probably consider to do REMD in those bad windows. This technique might work, provided you don't destabilize the membrane, but if the peptide is truly stuck in this orientation, I doubt that limited-range REMD would be very useful. -Justin Cheers Jianguo *From:* Justin A. Lemkul jalem...@vt.edu *To:* Gromacs Users' List gmx-users@gromacs.org *Sent:* Tuesday, 22 February 2011 21:10:08 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thanks Justin and Chris and sorry for confusing interpretation. Let me make it more clear. My peptide is flexible Martini beads, and highly positively charged. My membrane is a mixture of negatively charged lipids (25%) and zitterionic lipids(75%). So there is strong electrostatic attraction between peptide and membrane. To get the PMF, I did the following: (1) I did pulling simulation along (0 0 -1) direction to pull my peptide across the membrane. Then I got different configurations corresponding to different windows along the reaction coordinates, which is the z-distance between peptide and membrane. This figure (http://www.flickr.com/photos/lijg/5467080971/) shows some of the configurations at certain reaction coordinates. Are you not sampling configurations outside of the membrane (i.e. in water)? I would think that would solve your problem. You don't show any configurations in which the peptide is completely dissociated from the membrane. I don't know your objectives, but I would think that if you could completely extract the peptide from the membrane after passing through it, this would solve your problem. (2) In each window, I used the corresponding configuration that generated by the pulling simulation as initial input and run umbrella sampling. The size of each window is 0.15 nm, but close to the bilayer cneter (e.g., -0.6d0.6), I have increased number of windows so that the width of the window is to be 0.05 or 0.1 nm, I also tried to use different force constant in these windows. From the figure (http://www.flickr.com/photos/lijg/5467080971/) , we can classify the peptide conformation to be either extended (interacting with two bilayers) or compact (interacting with only one bilayer). Ideally, the peptide conformation should be similar for d=x and d=-x. The problem is that the configuration of peptide is not symmetric with respect to the bilayer center. For example, the peptide configuration is compact at d=0.6 and d=0.9, but the peptide is extended at d=-0.6 and d=-0.9. This leads Hysteresis. If I use g_wham to generate PMF, then the PMF is not symmetric with respect to the bilayer center. Using more number of windows and different force constant did not remove the problem. In my opinion, at least in some windows, the peptide should sample both compact and extended structure. But what I found is that the windowed Don't pre-judge the model :) Also, as I said before, there is no reason to suspect that MARTINI will produce any meaningful secondary structure changes. It was not parameterized to do so. umbrella simulation depends on the initial peptide conformation. If the initial peptide conformation is compact, then after 100 ns, it is still compact; if the initial peptide in that window is extended, the final configuration is also extended. I also tried to run longer equilibrium time (e.g., 200 ns), but the problem still exists. Sounds like a limitation of the force field model. My question is how to increase sampling of the peptide conformation? I just think of two choices: (1) use high temperature (e.g., 500K) at those bad windows. As I mentioned, I am wondering if g_wham can unbias the effect of using different temperatures
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Sorry, why do you think the PMF should be asymmetric? I pulled my peptide from d=9nm (above the membrane) to d=-3nm (below the membrane) and I did windowed umbrella sampling in the range of d=-1.05nm to d=9nm. At least the PMF should be symmetric with respect of the bilayer center in the range of d=[-1.05nm 1.05nm], something like a guassian distribution. But I got asymmetric PMF in this region. I also did reverse pulling starting from the peptide below the membrane ending with the peptide above the membrane. And the subsequent PMF of reversed pulling is also asymmetic. I have position restrains of the phosphate beads of the lipids in z-direction. So the membrane should be stable in REMD. But as you mentioned, if peptide is truly stuck in this orientation, REMD may be not useful. I will do a single simulation first at a higher temperature (e.g., 400K) in those bad windows to see if the peptide conformations are fully sampled. Cheers, Jianguo From: Justin A. Lemkul jalem...@vt.edu To: Gromacs Users' List gmx-users@gromacs.org Sent: Wednesday, 23 February 2011 10:24:46 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thank you, Justin. Actually I did windowed umbrella simulations from d=-1.05nm to d=9nm. Since I think there is no problem in the region out of the membrane, so I only show the configurations within the membrane. My objective is to access the free energy barrier of the peptide translocate the negatively charged membrane. The problem is that the PMF is not symmetric with respect to the bilayer center due to the unconverged simulations. I would argue that the PMF is not symmetric because your reaction coordinate is not symmetric. How can you calculate a free energy of crossing a charged membrane when your peptide does not cross the membrane? What I proposed earlier was to obtain configurations at equal distances above and below the membrane (arbitrary in a periodic system, but hopefully you get the idea). If you can extract the peptide to the point where it is liberated from the membrane in the negative direction, I'd suspect you could solve your problem. Since g_wham does not support different temperatures in different windows, to increase the converges, I will probably consider to do REMD in those bad windows. This technique might work, provided you don't destabilize the membrane, but if the peptide is truly stuck in this orientation, I doubt that limited-range REMD would be very useful. -Justin Cheers Jianguo *From:* Justin A. Lemkul jalem...@vt.edu *To:* Gromacs Users' List gmx-users@gromacs.org *Sent:* Tuesday, 22 February 2011 21:10:08 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thanks Justin and Chris and sorry for confusing interpretation. Let me make it more clear. My peptide is flexible Martini beads, and highly positively charged. My membrane is a mixture of negatively charged lipids (25%) and zitterionic lipids(75%). So there is strong electrostatic attraction between peptide and membrane. To get the PMF, I did the following: (1) I did pulling simulation along (0 0 -1) direction to pull my peptide across the membrane. Then I got different configurations corresponding to different windows along the reaction coordinates, which is the z-distance between peptide and membrane. This figure (http://www.flickr.com/photos/lijg/5467080971/) shows some of the configurations at certain reaction coordinates. Are you not sampling configurations outside of the membrane (i.e. in water)? I would think that would solve your problem. You don't show any configurations in which the peptide is completely dissociated from the membrane. I don't know your objectives, but I would think that if you could completely extract the peptide from the membrane after passing through it, this would solve your problem. (2) In each window, I used the corresponding configuration that generated by the pulling simulation as initial input and run umbrella sampling. The size of each window is 0.15 nm, but close to the bilayer cneter (e.g., -0.6d0.6), I have increased number of windows so that the width of the window is to be 0.05 or 0.1 nm, I also tried to use different force constant in these windows. From the figure (http://www.flickr.com/photos/lijg/5467080971/) , we can classify the peptide conformation to be either extended (interacting with two bilayers) or compact (interacting with only one bilayer). Ideally, the peptide conformation should be similar for d=x and d=-x. The problem is that the configuration of peptide is not symmetric with respect to the bilayer center. For example, the peptide configuration is compact at d=0.6
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Jianguo Li wrote: Dear all, I want to get the PMF of my peptide across the membrane bilayer. First I pulled my peptide across the membrane and then did windowed umbrella sampling along the reaction coordinates which is the z-distance between peptide and membrane. However, I found that sampling is not sufficient in some windows(e.g., around the center of the membrane). To enhance the sampling, I am thinking to run the simulation in those windows at higher temperature (e.g., 500K), but this will introduce a bias. My question is: can g_wham remove the bias due to using different temperatures in different windows? If g_wham cannot deal with the bias due to using different T, I may need to do REMD in those windows. But that will be very expensive computationally. Anybody have an idea of enhancing sampling in those windows? Btw, I am using Martini CG model. Any suggestions will be highly appreciated, thank you! A more straightforward approach is to (1) add more sampling windows or (2) increase the force constant in regions where there's poor sampling, or perhaps both. -Justin Cheers, Jianguo -- Justin A. Lemkul Ph.D. Candidate ICTAS Doctoral Scholar MILES-IGERT Trainee Department of Biochemistry Virginia Tech Blacksburg, VA jalemkul[at]vt.edu | (540) 231-9080 http://www.bevanlab.biochem.vt.edu/Pages/Personal/justin -- gmx-users mailing listgmx-users@gromacs.org http://lists.gromacs.org/mailman/listinfo/gmx-users Please search the archive at http://www.gromacs.org/Support/Mailing_Lists/Search before posting! Please don't post (un)subscribe requests to the list. Use the www interface or send it to gmx-users-requ...@gromacs.org. Can't post? Read http://www.gromacs.org/Support/Mailing_Lists
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Jianguo Li wrote: Thanks Justin. I tried your suggestions by either increase more windows and change the force constant, but it seems the samplings are still bad in some windows. When I did pulling in (0 0 1) direction and a reverse pulling in (0 0 -1) direction, I got different configurations at certain reaction coordinates. And the windowed umbrella sampling seems depends strongly on the initial configurations in that window. Therefore I got different PMFs using pulling in (0 0 1) direction and reverse pulling in （0 0 -1) direction. How long are each of the simulations in each window? Sufficient sampling should eliminate any configurational bias and/or hysteresis. Also, if the pulling that sets up the initial configurations is done slowly enough, you won't see these problems. Sounds to me like you're pulling too fast or hard, such that the system is not stable. In my simulation, I exert constraints on phosphate atoms in z direction, so there is no lipid flip-flop and the membrane will be stable at high temperatures. Then I am thinking of increasing temperature in those bad windows to enhance sampling... I don't know if I can make a convincing argument here, but intuitively, these windows would be sampling in a different ensemble, so the free energy landscape in these windows would be discontinuous with any adjacent windows that are done at different temperatures, and perhaps the forces required to restrain your peptide at a given COM distance will still result in a discontinuous PMF. I would also suspect that g_wham can't handle this situation; it has a -temp flag, but it only takes one value. So if you construct your PMF curve using WHAM, but supply incorrect or inconsistent information, I certainly wouldn't believe the result. I guess the main point is, there are tons of published demonstrations of peptides and other molecules crossing a membrane with SMD and umbrella sampling, so it should be possible to generate stable configurations without any funny tricks. -Justin best regards, Jianguo *From:* Justin A. Lemkul jalem...@vt.edu *To:* Discussion list for GROMACS users gmx-users@gromacs.org *Sent:* Tuesday, 22 February 2011 09:35:37 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Dear all, I want to get the PMF of my peptide across the membrane bilayer. First I pulled my peptide across the membrane and then did windowed umbrella sampling along the reaction coordinates which is the z-distance between peptide and membrane. However, I found that sampling is not sufficient in some windows(e.g., around the center of the membrane). To enhance the sampling, I am thinking to run the simulation in those windows at higher temperature (e.g., 500K), but this will introduce a bias. My question is: can g_wham remove the bias due to using different temperatures in different windows? If g_wham cannot deal with the bias due to using different T, I may need to do REMD in those windows. But that will be very expensive computationally. Anybody have an idea of enhancing sampling in those windows? Btw, I am using Martini CG model. Any suggestions will be highly appreciated, thank you! A more straightforward approach is to (1) add more sampling windows or (2) increase the force constant in regions where there's poor sampling, or perhaps both. -Justin Cheers, Jianguo -- Justin A. Lemkul Ph.D. Candidate ICTAS Doctoral Scholar MILES-IGERT Trainee Department of Biochemistry Virginia Tech Blacksburg, VA jalemkul[at]vt.edu | (540) 231-9080 http://www.bevanlab.biochem.vt.edu/Pages/Personal/justin -- gmx-users mailing listgmx-users@gromacs.org mailto:gmx-users@gromacs.org http://lists.gromacs.org/mailman/listinfo/gmx-users Please search the archive at http://www.gromacs.org/Support/Mailing_Lists/Search before posting! Please don't post (un)subscribe requests to the list. Use the www interface or send it to gmx-users-requ...@gromacs.org mailto:gmx-users-requ...@gromacs.org. Can't post? Read http://www.gromacs.org/Support/Mailing_Lists -- Justin A. Lemkul Ph.D. Candidate ICTAS Doctoral Scholar MILES-IGERT Trainee Department of Biochemistry Virginia Tech Blacksburg, VA jalemkul[at]vt.edu | (540) 231-9080 http://www.bevanlab.biochem.vt.edu/Pages/Personal/justin -- gmx-users mailing listgmx-users@gromacs.org http://lists.gromacs.org/mailman/listinfo/gmx-users Please search the archive at http://www.gromacs.org/Support/Mailing_Lists/Search before posting! Please don't post (un)subscribe requests to the list. Use the www interface or send it to gmx-users-requ...@gromacs.org. Can't post? Read http://www.gromacs.org
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Thanks for your comments, Justin. Using timestep of 20 fs, in each window the simulation runs for 100 ns CG time. The pulling rate is 0.001 nm/ps. Is it too fast? My system is a little different. My peptide is highly positively charged. The NMR experiments show that the conformation of the peptide in water is very dynamic, so I make it flexible without fixing any secondary structure in Martini model. In the membrane, 25% of the lipids are negatively charged, so there are very strong electrostatic attraction between peptides and membrane. During the peptide approaching the membrane from the top, peptide can take different configurations at different reaction coordinates. When pulling the peptide into the membrane, the peptide takes relatively compact structure and interacts with only the top leaflet until the distance becomes smaller than 0.45 nm, after that the peptide becomes extended structure and interacts with both leaflets. This extended structure remains until the distance becomes -1.05 nm. Further pulling leads to compact structure and interacts only with the lower leaflet. So the comformation of the peptide is not symmetric between the center of the bilayer, which leads to Hysteresis. It seems that there is a huge energy barrier for the peptide to translocate across the membrane because if the initial conformation in a certain window is extended (interacting with both leaflets), then it remains extended. Similarly, it the initial conformation in a certain window is compact (interacting with only one leaflet), it will remain compact. Any Suggestions of dealing with the highly charged system? Cheers, Jianguo From: Justin A. Lemkul jalem...@vt.edu To: Gromacs Users' List gmx-users@gromacs.org Sent: Tuesday, 22 February 2011 09:58:36 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thanks Justin. I tried your suggestions by either increase more windows and change the force constant, but it seems the samplings are still bad in some windows. When I did pulling in (0 0 1) direction and a reverse pulling in (0 0 -1) direction, I got different configurations at certain reaction coordinates. And the windowed umbrella sampling seems depends strongly on the initial configurations in that window. Therefore I got different PMFs using pulling in (0 0 1) direction and reverse pulling in （0 0 -1) direction. How long are each of the simulations in each window? Sufficient sampling should eliminate any configurational bias and/or hysteresis. Also, if the pulling that sets up the initial configurations is done slowly enough, you won't see these problems. Sounds to me like you're pulling too fast or hard, such that the system is not stable. In my simulation, I exert constraints on phosphate atoms in z direction, so there is no lipid flip-flop and the membrane will be stable at high temperatures. Then I am thinking of increasing temperature in those bad windows to enhance sampling... I don't know if I can make a convincing argument here, but intuitively, these windows would be sampling in a different ensemble, so the free energy landscape in these windows would be discontinuous with any adjacent windows that are done at different temperatures, and perhaps the forces required to restrain your peptide at a given COM distance will still result in a discontinuous PMF. I would also suspect that g_wham can't handle this situation; it has a -temp flag, but it only takes one value. So if you construct your PMF curve using WHAM, but supply incorrect or inconsistent information, I certainly wouldn't believe the result. I guess the main point is, there are tons of published demonstrations of peptides and other molecules crossing a membrane with SMD and umbrella sampling, so it should be possible to generate stable configurations without any funny tricks. -Justin best regards, Jianguo *From:* Justin A. Lemkul jalem...@vt.edu *To:* Discussion list for GROMACS users gmx-users@gromacs.org *Sent:* Tuesday, 22 February 2011 09:35:37 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Dear all, I want to get the PMF of my peptide across the membrane bilayer. First I pulled my peptide across the membrane and then did windowed umbrella sampling along the reaction coordinates which is the z-distance between peptide and membrane. However, I found that sampling is not sufficient in some windows(e.g., around the center of the membrane). To enhance the sampling, I am thinking to run the simulation in those windows at higher temperature (e.g., 500K), but this will introduce a bias. My question is: can g_wham remove the bias due to using different temperatures in different windows
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Jianguo Li wrote: Thanks for your comments, Justin. Using timestep of 20 fs, in each window the simulation runs for 100 ns CG time. The pulling rate is 0.001 nm/ps. Is it too fast? Let me clarify things, since I'm not convinced I understand your procedure. You generate a series of configurations with 0.001 nm/ps pulling, but then how many windows do you generate for independent simulations? What are your .mdp parameters during those windows? The pull rate should be 0 during the actual umbrella sampling, to restrain the peptide within the window. What force constant(s) do you use? My system is a little different. My peptide is highly positively charged. The NMR experiments show that the conformation of the peptide in water is very dynamic, so I make it flexible without fixing any secondary structure in Martini model. As was discussed in the last few days, do not interpret changes in structure too directly when using MARTINI. It is not designed to faithfully mimic secondary structure changes. In the membrane, 25% of the lipids are negatively charged, so there are very strong electrostatic attraction between peptides and membrane. During the peptide approaching the membrane from the top, peptide can take different configurations at different reaction coordinates. When pulling the peptide into the membrane, the peptide takes relatively compact structure and interacts with only the top leaflet until the distance becomes smaller than 0.45 nm, after that the peptide becomes extended structure and interacts with both leaflets. This extended structure remains until the distance becomes -1.05 nm. Further pulling leads to compact structure and interacts only with the lower leaflet. So the comformation of the peptide is not symmetric between the center of the bilayer, which leads to Hysteresis. It seems that there is a huge I guess I'm confused here, too. The peptide is compact when interacting with the top leaflet, extended further in the membrane, then compact again when interacting with the lower leaflet. What's strange about that? energy barrier for the peptide to translocate across the membrane because if the initial conformation in a certain window is extended (interacting with both leaflets), then it remains extended. Similarly, it the initial conformation in a certain window is compact (interacting with only one leaflet), it will remain compact. I don't see how that is necessarily unexpected or problematic. Peptides will change conformation depending on their environment. If you want a static structure to cross the membrane (which may or may not represent reality) you'll have to introduce some kind of intramolecular restraints. -Justin Any Suggestions of dealing with the highly charged system? Cheers, Jianguo *From:* Justin A. Lemkul jalem...@vt.edu *To:* Gromacs Users' List gmx-users@gromacs.org *Sent:* Tuesday, 22 February 2011 09:58:36 *Subject:* Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thanks Justin. I tried your suggestions by either increase more windows and change the force constant, but it seems the samplings are still bad in some windows. When I did pulling in (0 0 1) direction and a reverse pulling in (0 0 -1) direction, I got different configurations at certain reaction coordinates. And the windowed umbrella sampling seems depends strongly on the initial configurations in that window. Therefore I got different PMFs using pulling in (0 0 1) direction and reverse pulling in （0 0 -1) direction. How long are each of the simulations in each window? Sufficient sampling should eliminate any configurational bias and/or hysteresis. Also, if the pulling that sets up the initial configurations is done slowly enough, you won't see these problems. Sounds to me like you're pulling too fast or hard, such that the system is not stable. In my simulation, I exert constraints on phosphate atoms in z direction, so there is no lipid flip-flop and the membrane will be stable at high temperatures. Then I am thinking of increasing temperature in those bad windows to enhance sampling... I don't know if I can make a convincing argument here, but intuitively, these windows would be sampling in a different ensemble, so the free energy landscape in these windows would be discontinuous with any adjacent windows that are done at different temperatures, and perhaps the forces required to restrain your peptide at a given COM distance will still result in a discontinuous PMF. I would also suspect that g_wham can't handle this situation; it has a -temp flag, but it only takes one value. So if you construct your PMF curve using WHAM, but supply incorrect or inconsistent information, I certainly wouldn't believe the result. I guess the main point is, there are tons of published
Re: [gmx-users] Can g_wham support using different temperature for different windows?
Thanks Justin and Chris and sorry for confusing interpretation. Let me make it more clear. My peptide is flexible Martini beads, and highly positively charged. My membrane is a mixture of negatively charged lipids (25%) and zitterionic lipids(75%). So there is strong electrostatic attraction between peptide and membrane. To get the PMF, I did the following: (1) I did pulling simulation along (0 0 -1) direction to pull my peptide across the membrane. Then I got different configurations corresponding to different windows along the reaction coordinates, which is the z-distance between peptide and membrane. This figure (http://www.flickr.com/photos/lijg/5467080971/) shows some of the configurations at certain reaction coordinates. (2) In each window, I used the corresponding configuration that generated by the pulling simulation as initial input and run umbrella sampling. The size of each window is 0.15 nm, but close to the bilayer cneter (e.g., -0.6d0.6), I have increased number of windows so that the width of the window is to be 0.05 or 0.1 nm, I also tried to use different force constant in these windows. From the figure (http://www.flickr.com/photos/lijg/5467080971/) , we can classify the peptide conformation to be either extended (interacting with two bilayers) or compact (interacting with only one bilayer). Ideally, the peptide conformation should be similar for d=x and d=-x. The problem is that the configuration of peptide is not symmetric with respect to the bilayer center. For example, the peptide configuration is compact at d=0.6 and d=0.9, but the peptide is extended at d=-0.6 and d=-0.9. This leads Hysteresis. If I use g_wham to generate PMF, then the PMF is not symmetric with respect to the bilayer center. Using more number of windows and different force constant did not remove the problem. In my opinion, at least in some windows, the peptide should sample both compact and extended structure. But what I found is that the windowed umbrella simulation depends on the initial peptide conformation. If the initial peptide conformation is compact, then after 100 ns, it is still compact; if the initial peptide in that window is extended, the final configuration is also extended. I also tried to run longer equilibrium time (e.g., 200 ns), but the problem still exists. My question is how to increase sampling of the peptide conformation? I just think of two choices: (1) use high temperature (e.g., 500K) at those bad windows. As I mentioned, I am wondering if g_wham can unbias the effect of using different temperatures in different windows. (2) use REMD in those bad windows. These need a lot of computational resources. Is there any other method to deal with the insufficient sampling? Any suggestions are welcome, thanks for your time reading this email! Cheers, Jianguo From: Justin A. Lemkul jalem...@vt.edu To: Gromacs Users' List gmx-users@gromacs.org Sent: Tuesday, 22 February 2011 11:13:05 Subject: Re: [gmx-users] Can g_wham support using different temperature for different windows? Jianguo Li wrote: Thanks for your comments, Justin. Using timestep of 20 fs, in each window the simulation runs for 100 ns CG time. The pulling rate is 0.001 nm/ps. Is it too fast? Let me clarify things, since I'm not convinced I understand your procedure. You generate a series of configurations with 0.001 nm/ps pulling, but then how many windows do you generate for independent simulations? What are your .mdp parameters during those windows? The pull rate should be 0 during the actual umbrella sampling, to restrain the peptide within the window. What force constant(s) do you use? My system is a little different. My peptide is highly positively charged. The NMR experiments show that the conformation of the peptide in water is very dynamic, so I make it flexible without fixing any secondary structure in Martini model. As was discussed in the last few days, do not interpret changes in structure too directly when using MARTINI. It is not designed to faithfully mimic secondary structure changes. In the membrane, 25% of the lipids are negatively charged, so there are very strong electrostatic attraction between peptides and membrane. During the peptide approaching the membrane from the top, peptide can take different configurations at different reaction coordinates. When pulling the peptide into the membrane, the peptide takes relatively compact structure and interacts with only the top leaflet until the distance becomes smaller than 0.45 nm, after that the peptide becomes extended structure and interacts with both leaflets. This extended structure remains until the distance becomes -1.05 nm. Further pulling leads to compact structure and interacts only with the lower leaflet. So the comformation of the peptide is not symmetric between the center of the bilayer, which leads to Hysteresis. It seems
