Wednesday, July 4, 2018

NMRlipids IV: First draft of the manuscript about PS lipids

Thank you again for the 168 comments in the two previous blog posts about NMRlipids IV project (NMRlipids IV: Headgroup & glycerol backbone structures, and cation binding in bilayers with PE, PG and PS lipids and NMRlipids IV: Current status and reorganization of the manuscript) and numerous contributions to the GitHub repository.

First draft of the manuscript summarizing the results related to PS lipids is now compiled. As discussed previously, the goal is to first finish the manuscript about PS headgroup and then progress with PG and PE results separately. In addition to the up-to-date manuscript (pdf, tex) and supplementary information (pdf, tex), also the figures and data are available in the GitHub repository.

I believe that the manuscript already contains most of our conclusions, but several issues need to be addressed before we can start to prepare the submission. Most important current issues are listed here, accompanied with the names of persons who can hopefully help me to resolve the issues.
  1. We need slightly more detailed description of the NMR experiments by Tiago Ferreira. Also the results and discussion about the experiments, including Figure 1, needs to be polished.
  2. Atom names and dihedral notations should be made consistent between the chemical structures in figure 2, and dihedral angles distributions in  figures S6 and S7 by Pavel Buslaev. I am not sure if we could just the atom names currently used in figure 2, or if we need more labels. Slight polishing of figures S6 and S7 is also needed.
  3. The subjective ranking criteria in figure 4 by Markus Miettinen should be probably improved, see issue #4 in the GitHub repository.
  4. CHARMM36 data from POPC:POPS (5:1) mixtures with added NaCl would be highly useful for figure 6. CHARMM36 gives two maxima in the counterion density profiles in figure 5, while for example Lipid17 gives only one. From the right column of figure 6 we could see if this difference is reflected to the order parameter response of PS headgroup to the excess counterion concentration. Data from Gromos-CKP would also be useful for this. If someone has such data or is willing to generate it, let us know. 
  5. CHARMM36 simulations with the added CaCl2, but without the new NBfix, mentioned by Jesper Madsen would be useful for figure 9
  6. Lipid17 simulations ran with the Amber package, mentioned by Batuhan Kav, would be useful for figure 9
  7. Details of simulations by Thomas Piggot, Jesper Madsen, Fernando Favela, Batuhan Kav, Markus Miettinen, Josef Melcr and Matti Javanainen should be added in section S1 in the supplementary information (tex file here).
In addition, all comments regarding the current manuscript are welcomed. I will continue working with the manuscript and keep the the list updated.  


  1. Regarding the issue number 4 in the above list, I realized that the mentioned data for Gromos-CKP has already been contributed by Tom Piggot:

    I will proceed in adding this into the manuscript.

    1. I have now added the mentioned data into the right column of figure 8 ( However, for the left column we would need the pure "GROMOS-CKP POPC" simulation. Which force field was used for POPC in the simulations of mixtures?

      In addition, simulations of the POPC:POPS mixtures with additional NaCl concentration would be useful for figure 6 (

  2. Regarding figures S6-S8, please find updated versions here:

    I have also shared the figure with atom labels. If anything is wrong with the figures or labels, please tell me, I will correct everything

    1. Thanks a lot. I think that the figS8.png is highly useful and I incorporated it into the main manuscript.

      Here are some comments:
      1) We (and most lipid people) label the glycerol backbone carbon with the headgroup attached as g_3, and the carbon where sn-1 chain is attached as g_1. Therefore, the 1 and 3 labels should be swapped your current labeling.
      2) The analysis of MacRog may still be done from the system which was accidentally OPPS, not POPS (see The data for MacRog POPS is available from
      3) It would be useful to have a list of simulations used in the analysis. Also the analysis codes/scripts might be useful.

    2. Dear Samuli,
      I have reworked the figures for the new labeling and correct MacRog trajectory. Please find new versions here:

      For the analysis I used the trajectories POPS/Na+ for CHARMM36, CHARMM36UA, Slipids, Berger, MacRog, GROMOS-CKPM, GROMOS-CKP, Lipid17/JC, Lipid17/ff99 from Table1 of the manuscript.

    3. Thanks again. Now I am comparing the new and previous versions of the figS8.png.

      The orientation of the headgroup snapshots from MacRog in the previous version seems to be somehow different than in other models:

      In the new version the orientation seems to be similar to the other models:

      In addition, the headgroup dihedral distributions from MacRog in the new and previous plots seems to be different, even thought the force field should be identical in this region:

      Maybe Tom Piggot or Matti Javanainen could also have some insight in this?

    4. I think this could well be due to the use of different starting structures in these two simulations (like that seen previously with the OPLS-UA Ulmschneider PC simulations). My simulations with the PO tails were started from the same membrane structure as all of other POPS simulations I did. This structure was originally a CHARMM36 POPS membrane made using the CHARMM-GUI.

    5. I will double check the two topologies to check there isn't anything different here but I'm pretty confident there won't be

    6. Matti Javanainen gave me a preliminary trajectory from POPC:POPS (5:1) simulation with the fixed itp file by Tom Piggot. I compared the POPS order parameter results to the ones from POPC:OPPS (5:1):

      The results are quite similar, in contrast to the previous comparison of pure POPS simulations which gave significant differences between the models:

      This suggested that the problem would be in the initial conformations, as also indicated by Tom Piggot above. Therefore, we took a look to initial configurations with Matti Javanainen and noticed that the starting configuration shared by the MacRog developers has the serine in D-stereoisomer,
      while other models have the L-stereoisomer. The latter is present in biological systems.

    7. I have rewritten the discussing about conformations and dihedrals, and pushed it into the current manuscript in GitHub. For the headgroup, it is now proposed that the rigid C_alpha-C_beta-C_gamma-O_gamma dihedral in CHARMM36 and Slipid is more realistic than more flexible structures in other models. There is also some discussion about N-C_beta-C_aplha-O_alpha dihedral. Conclusions from the glycerol backbone would require some more elaborate analysis.

      I still have few requests about the pictures:
      1) Could you swap the order in figS8.png such that the headgroup structures would be on top and glycerol backbone structures on bottom? This would be more fluent with the discussion in the manuscript.
      2) There is a typo in the labeling of "Brger" in S6 and S7.
      3) GROMOS-CKP2 model is labeled twice. Note that the current naming convention of CKP models is described in the caption of Table 1.
      4) N-C_beta-C_alpha-O_alpha dihedral seems very similar for CHARMM36 and CHARMM36ua in the figS8.png. However, maximum at 60 degrees seems to be missing from the CHARMM36ua results in Fig. S6. Do you have any idea what could be the reason for this?
      5) Higher resolution version of figure PS_Labels.png would be useful.

    8. Dear Samuli,
      I have reworked the figures as you asked. As for the question 4 - there was a problem with my script. Now everything is ok. Please, find new figures here:

    9. Dear Pavel,
      thanks again for the update. I have now included the new figures and updated the discussion in the manuscript.

      Two more questions:
      1) The dihedral distributions from CHARMM36 and CHARMM36UA seem to overlap almost perfectly. Is this correct? If yes, maybe the line for CHARMM36 should be slightly thicker to make it visible. Now it is quite difficult to distinguish from some figures
      2) Would it be possible to make similar figures (snapshots and headgroup dihedral distributions) from CHARMM36 and Slipids POPC trajectories, available from and, respectively? I think that the differences between headgroup order parameters of POPC and POPS are roughly reproduced by these models, thus we could consider using these to look at the structural differences.

    10. Dear Samuli,
      1. I have reworked figures S6 and S7. Now both lines seem to be visible. Please find them via link:
      2. Regarding the POPC trajectories, I will do this during this week. Hopefully, by Tuesday.

    11. Dear Samuli,
      I have plotted figures analogous to S6 and S7 for POPC lipids. Please find them via link (figS6PC, figS7PC):

      There are some differences in distributions for PC and PS headgroups. Thus, I will plot structures soon.

    12. Thanks for the results.

      Should the label for the green dashed line be "POPC, Slipids"?

      The dashed lines in the figS6PC are the same for each dihedral.

    13. I have changed the names.

      For the dashed lines, I did not account for different number of calculated angles for PC and PS lipids. Now should be ok.

      Please find plots via link:

    14. Dear Samuli,

      I have added structural figures for glycerol backbone (figS8_POPC at It represents the differences in the distributions (figS6PC) between Slipids POPC and POPS lipids.

    15. Thanks. Could you also make a figure about the headgroup part, i.e., from the figure presenting the figS7PC? This would be actually more interesting when comparing PC and PS.

    16. I have added the figures for the headgroup to. Please find it via link:

    17. I have now composed a new figure summarizing the most essential conformations for the manuscript and included this into the manuscript and updated the discussion:

      I think that we still need a figure with atom labels for the b) part of this figure. Would it be possible to make similar labelling figure for b) as in the part a)? Maybe it would be also possible to have this kind of figure with high resolution?

      Other figures are now in the SI but are referred in the text.

    18. Dear Samuli, sorry for the late reply.

      Please find labeling figures via link:

    19. Thanks again. I made a new version of the figure:

      The atom labels in the left column are now too small. Could you make new versions of figures titled chol.png and pc_hg.png in your drive having atom labels with larger font?

    20. Dear Samuli, I have updated the figures. Are they good now?

  3. I have now started simulations of POPC:POPS (5:1) mixture with additional amount of NaCl and KCl counterions, which were requested in todo point 4.

    1. These results are now added in the figure:

      There is also a new post discussing about the evaluation of counterion binding affinity to PS containing bilayers:

  4. I generated starting configurations for the PC/PS simulations using CHARMM-GUI and rerun the mixtures with KCl. These simulations should have the correct stereoisomer. The new data are included in the old Zenodo DOI: 10.5281/zenodo.1404040. The data with CaCl_2 will follow once the ion binding has converged. The salt-free system will also be included in the CaCl_2 upload.

    1. And here are the data for the same membranes with CaCl2.

    2. These are now included in the manuscript.


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