# Please how can i neutralize the system?

**URL:** <https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757>\
**Category:** User discussions\
**Tags:** mdp-parameters\
**Created:** [September 1, 2021, 8:19pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757 "2021-09-01T20:19:33Z")\
**Posts on this page:** 11\
**Page:** 1

<div class="post-metadata">

**Author:** ![serta](https://avatars.discourse-cdn.com/v4/letter/s/65b543/32.png) [@serta](https://gromacs.bioexcel.eu/u/serta)\
**Post date:** [September 1, 2021, 8:19pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/1 "2021-09-01T20:19:33Z")

</div>

GROMACS version:2021  
GROMACS modification: No  
Here post your question

hello, i ve got this problem during Md run; please how can i neutralize the system?  
when i add NA it will be 0.66000.

```auto
*NOTE 2 [file [topol.top](http://topol.top/), line 56]:*
*System has non-zero total charge: -0.340000*
*Total charge should normally be an integer. See*
*http://www.gromacs.org/Documentation/Floating_Point_Arithmetic*
*for discussion on how close it should be to an integer.*

*WARNING 1 [file [topol.top](http://topol.top/), line 56]:*
*You are using Ewald electrostatics in a system with net charge. This can*
*lead to severe artifacts, such as ions moving into regions with low*
*dielectric, due to the uniform background charge. We suggest to*
*neutralize your system with counter ions, possibly in combination with a*
*physiological salt concentration.*

```

I can pass this problem using -maxwarn in command line  
or to change this line in mdp files:  
coulombtype = PME ; Particle Mesh Ewald for long-range electrostatics  
to  
coulombtype = cutoff ; Particle Mesh Ewald for long-range electrostatics

Is it normal to use those solutions during MD? if not what is the solution to avoid the error?

---

<div class="post-metadata">

**Author:** ![jalemkul](https://dub1.discourse-cdn.com/flex017/user_avatar/gromacs.bioexcel.eu/jalemkul/32/18_2.png) [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)\
**Post date:** [September 1, 2021, 8:43pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/2 "2021-09-01T20:43:12Z")

</div>

A fractional charge of that magnitude means your topology is broken. -0.34 net charge is impossible.

---

<div class="post-metadata">

**Author:** ![serta](https://avatars.discourse-cdn.com/v4/letter/s/65b543/32.png) [@serta](https://gromacs.bioexcel.eu/u/serta)\
**Post date:** [September 1, 2021, 8:52pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/3 "2021-09-01T20:52:34Z")

</div>

how can i fix it?

---

<div class="post-metadata">

**Author:** ![jalemkul](https://dub1.discourse-cdn.com/flex017/user_avatar/gromacs.bioexcel.eu/jalemkul/32/18_2.png) [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)\
**Post date:** [September 1, 2021, 9:11pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/4 "2021-09-01T21:11:54Z")

</div>

You need to tell us what you’ve done and what is in your system. Provide your entire `pdb2gmx` command and the full screen output from it.

---

<div class="post-metadata">

**Author:** ![serta](https://avatars.discourse-cdn.com/v4/letter/s/65b543/32.png) [@serta](https://gromacs.bioexcel.eu/u/serta)\
**Post date:** [September 2, 2021, 9:27am UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/5 "2021-09-02T09:27:53Z")

</div>

Thank you for your response.  
I have used this lines; and my solvent system is water

> gmx editconf -f complex.gro -o newbox.gro -bt dodecahedron -d 1.0

 ![1](https://europe1.discourse-cdn.com/flex017/uploads/bioexcel1/original/2X/f/f0dcd6005ff0c34d79b8afeaa2fa0f5bbcb43b9f.png)

> gmx solvate -cp newbox.gro -cs spc216.gro -p topol.top -o solv.gro

 ![2](https://europe1.discourse-cdn.com/flex017/uploads/bioexcel1/original/2X/2/2aa70399e307b2d92d0a4f65737ec621b0024a39.png)

we have used the file of solvant ions from [Protein-Ligand Complex](http://www.mdtutorials.com/gmx/complex/04_ions.html)

> gmx grompp -f ions.mdp -c solv.gro -p topol.top -o ions.tpr

 ![3](https://europe1.discourse-cdn.com/flex017/uploads/bioexcel1/original/2X/2/26937356604d61c8b58a4b8255f8a3686bfc19f8.png)

> gmx genion -s ions.tpr -o solv\_ions.gro -p topol.top -pname NA -nname CL -neutral

 ![4](https://europe1.discourse-cdn.com/flex017/uploads/bioexcel1/original/2X/5/5edce329040d6833f757fc1e4cae5074f841bbbf.png)

and for Energy Minimization

we have used the file of solvent ions from open this: [http://www.mdtutorials.com/gmx/complex/Files/em.mdp](http://www.mdtutorials.com/gmx/complex/Files/em.mdp)

> gmx grompp -f em.mdp -c solv\_ions.gro -p topol.top -o em.tpr

this shows the error mentioned in the topic

---

<div class="post-metadata">

**Author:** ![jalemkul](https://dub1.discourse-cdn.com/flex017/user_avatar/gromacs.bioexcel.eu/jalemkul/32/18_2.png) [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)\
**Post date:** [September 2, 2021, 9:57am UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/6 "2021-09-02T09:57:24Z")

</div>

The problem comes from `pdb2gmx` but you did not provide that information. That is where the problem is. We need the full command and screen output (copy and paste is better than screenshots).

---

<div class="post-metadata">

**Author:** ![serta](https://avatars.discourse-cdn.com/v4/letter/s/65b543/32.png) [@serta](https://gromacs.bioexcel.eu/u/serta)\
**Post date:** [September 2, 2021, 1:45pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/7 "2021-09-02T13:45:54Z")

</div>

I have this command:

` gmx pdb2gmx -f 5fsa6139_clean.pdb -o 5fsa6139_processed.gro`

> gmx pdb2gmx -f 5fsah\_clean.pdb -o 5fsah\_processed.gro
> 
> Select the Force Field:  
> From current directory:  
> 1: CHARMM36 all-atom force field (July 2020)  
> From ‘/usr/share/gromacs/top’:  
> 2: AMBER03 protein, nucleic AMBER94 (Duan et al., J. Comp. Chem. 24, 1999-2012, 2003)  
> 3: AMBER94 force field (Cornell et al., JACS 117, 5179-5197, 1995)  
> 4: AMBER96 protein, nucleic AMBER94 (Kollman et al., Acc. Chem. Res. 29, 461-469, 1996)  
> 5: AMBER99 protein, nucleic AMBER94 (Wang et al., J. Comp. Chem. 21, 1049-1074, 2000)  
> 6: AMBER99SB protein, nucleic AMBER94 (Hornak et al., Proteins 65, 712-725, 2006)  
> 7: AMBER99SB-ILDN protein, nucleic AMBER94 (Lindorff-Larsen et al., Proteins 78, 1950-58, 2010)  
> 8: AMBERGS force field (Garcia & Sanbonmatsu, PNAS 99, 2782-2787, 2002)  
> 9: CHARMM27 all-atom force field (CHARM22 plus CMAP for proteins)  
> 10: GROMOS96 43a1 force field  
> 11: GROMOS96 43a2 force field (improved alkane dihedrals)  
> 12: GROMOS96 45a3 force field (Schuler JCC 2001 22 1205)  
> 13: GROMOS96 53a5 force field (JCC 2004 vol 25 pag 1656)  
> 14: GROMOS96 53a6 force field (JCC 2004 vol 25 pag 1656)  
> 15: GROMOS96 54a7 force field (Eur. Biophys. J. (2011), 40, 843-856, DOI: 10.1007/s00249-011-0700-9)  
> 16: OPLS-AA/L all-atom force field (2001 aminoacid dihedrals)  
> 1
> 
> Using the Charmm36-feb2021 force field in directory ./charmm36-feb2021.ff
> 
> Opening force field file ./charmm36-feb2021.ff/watermodels.dat
> 
> Select the Water Model:  
> 1: TIP3P TIP 3-point, recommended, by default uses CHARMM TIP3 with LJ on H  
> 2: TIP4P TIP 4-point  
> 3: TIP5P TIP 5-point  
> 4: SPC simple point charge  
> 5: SPC/E extended simple point charge  
> 6: None  
> 1  
> going to rename ./charmm36-feb2021.ff/merged.r2b  
> Opening force field file ./charmm36-feb2021.ff/merged.r2b  
> Reading 5fsah\_clean.pdb…  
> WARNING: all CONECT records are ignored  
> Read ‘’, 4833 atoms  
> Analyzing pdb file  
> Splitting chemical chains based on TER records or chain id changing.  
> There are 2 chains and 0 blocks of water and 485 residues with 4833 atoms
> 
> chain #res #atoms  
> 1 ‘A’ 484 4790  
> 2 ’ ’ 1 43
> 
> WARNING: there were 4 atoms with zero occupancy and 34 atoms with  
> occupancy unequal to one (out of 4833 atoms). Check your pdb file.
> 
> Opening force field file ./charmm36-feb2021.ff/atomtypes.atp  
> Reading residue database… (Charmm36-feb2021)  
> Opening force field file ./charmm36-feb2021.ff/merged.rtp  
> Opening force field file ./charmm36-feb2021.ff/merged.hdb  
> Opening force field file ./charmm36-feb2021.ff/merged.n.tdb  
> Opening force field file ./charmm36-feb2021.ff/merged.c.tdb  
> Processing chain 1 ‘A’ (4790 atoms, 484 residues)  
> Analysing hydrogen-bonding network for automated assignment of histidine  
> protonation. 741 donors and 722 acceptors were found.  
> There are 956 hydrogen bonds  
> Will use HISE for residue 101  
> Will use HISE for residue 120  
> Will use HISE for residue 183  
> Will use HISE for residue 242  
> Will use HISE for residue 283  
> Will use HISE for residue 310  
> Will use HISE for residue 322  
> Will use HISE for residue 328  
> Will use HISE for residue 373  
> Will use HISE for residue 377  
> Will use HISE for residue 400  
> Will use HISE for residue 410  
> Will use HISE for residue 468  
> Identified residue LYS45 as a starting terminus.  
> Identified residue PHE528 as a ending terminus.  
> 8 out of 8 lines of specbond.dat converted successfully  
> Special Atom Distance matrix:  
> CYS75 MET86 MET92 HIS101 HIS120 CYS134 MET140  
> SG310 SD428 SD482 NE2565 NE2751 SG872 SD932  
> MET86 SD428 1.393  
> MET92 SD482 1.275 0.978  
> HIS101 NE2565 1.617 2.461 1.626  
> HIS120 NE2751 2.804 1.962 1.549 2.751  
> CYS134 SG872 3.417 3.052 2.266 2.599 1.448  
> MET140 SD932 3.160 3.184 2.267 1.966 2.149 1.036  
> HIS183 NE21394 4.841 5.294 4.811 4.537 4.983 5.035 5.130  
> MET189 SD1441 2.805 3.092 2.521 2.538 2.776 2.995 3.118  
> MET209 SD1633 4.710 4.555 3.805 3.842 2.988 2.323 2.826  
> HIS242 NE21971 3.071 2.596 2.227 3.090 1.650 2.370 2.984  
> MET258 SD2149 4.631 4.160 3.503 3.913 2.295 1.679 2.528  
> HIS283 NE22415 4.741 4.771 3.855 3.411 3.307 1.915 1.797  
> MET292 SD2503 4.112 3.867 3.039 3.084 2.206 0.872 1.365  
> MET306 SD2628 3.288 3.073 2.375 2.733 1.763 1.682 2.211  
> HIS310 NE22663 2.917 2.882 2.181 2.425 1.957 2.017 2.341  
> HIS322 NE22776 2.430 3.303 2.689 2.005 3.528 3.578 3.297  
> HIS328 NE22833 3.806 4.869 4.212 3.091 5.040 4.785 4.312  
> MET372 SD3268 2.230 3.229 2.594 1.720 3.566 3.575 3.190  
> HIS373 NE23281 2.217 3.041 2.399 1.772 3.237 3.305 3.039  
> MET374 SD3290 1.898 2.572 1.684 0.776 2.524 2.353 1.910  
> HIS377 NE23319 1.495 1.796 1.101 1.460 1.974 2.418 2.384  
> HIS400 NE23554 1.577 0.998 0.966 2.124 1.967 2.687 2.624  
> HIS410 NE23646 1.936 2.815 1.967 0.629 2.939 2.706 2.121  
> HIS468 NE24198 2.583 2.544 1.629 1.611 1.693 1.060 0.641  
> CYS470 SG4223 2.816 3.157 2.197 1.549 2.390 1.693 1.286  
> MET508 SD4594 2.306 2.165 1.455 2.015 1.443 1.822 2.111  
> CYS526 SG4769 5.803 6.354 5.672 5.038 5.738 5.351 5.308  
> MET527 SD4777 5.053 5.597 4.857 4.183 4.933 4.477 4.392  
> HIS183 MET189 MET209 HIS242 MET258 HIS283 MET292  
> NE21394 SD1441 SD1633 NE21971 SD2149 NE22415 SD2503  
> MET189 SD1441 2.312  
> MET209 SD1633 3.661 2.624  
> HIS242 NE21971 3.516 1.601 2.401  
> MET258 SD2149 4.654 3.155 1.237 2.313  
> HIS283 NE22415 5.075 3.669 2.069 3.631 2.114  
> MET292 SD2503 4.943 3.191 1.795 2.730 1.294 1.167  
> MET306 SD2628 3.469 1.581 1.510 1.099 1.614 2.566 1.800  
> HIS310 NE22663 3.107 1.043 1.893 1.082 2.173 2.865 2.213  
> HIS322 NE22776 2.673 1.349 3.584 2.796 4.158 4.050 3.813  
> HIS328 NE22833 2.781 2.714 4.412 4.280 5.248 4.665 4.811  
> MET372 SD3268 3.063 1.679 3.795 3.017 4.309 4.056 3.847  
> HIS373 NE23281 2.839 1.231 3.439 2.578 3.942 3.869 3.577  
> MET374 SD3290 3.829 1.813 3.187 2.513 3.402 3.081 2.733  
> HIS377 NE23319 3.730 1.471 3.252 1.776 3.308 3.625 2.966  
> HIS400 NE23554 5.768 3.486 4.580 3.055 4.098 4.359 3.532  
> HIS410 NE23646 3.975 2.124 3.540 2.958 3.803 3.279 3.055  
> HIS468 NE24198 4.969 2.809 2.970 2.601 2.636 2.344 1.693  
> CYS470 SG4223 3.895 2.011 2.356 2.446 2.595 2.064 1.818  
> MET508 SD4594 3.651 1.376 2.475 1.091 2.446 3.090 2.298  
> CYS526 SG4769 1.703 3.287 3.652 4.488 4.854 4.752 5.006  
> MET527 SD4777 1.752 2.598 2.947 3.816 4.118 3.906 4.149  
> MET306 HIS310 HIS322 HIS328 MET372 HIS373 MET374  
> SD2628 NE22663 NE22776 NE22833 SD3268 NE23281 SD3290  
> HIS310 NE22663 0.565  
> HIS322 NE22776 2.624 2.081  
> HIS328 NE22833 3.903 3.426 1.637  
> MET372 SD3268 2.807 2.283 0.413 1.642  
> HIS373 NE23281 2.402 1.863 0.302 1.897 0.455  
> MET374 SD3290 2.089 1.725 1.478 2.720 1.318 1.212  
> HIS377 NE23319 1.836 1.425 1.664 3.251 1.664 1.372 1.050  
> HIS400 NE23554 3.215 3.100 3.546 4.993 3.388 3.265 2.452  
> HIS410 NE23646 2.525 2.153 1.478 2.515 1.209 1.268 0.458  
> HIS468 NE24198 2.024 2.076 3.006 4.205 2.898 2.724 1.584  
> CYS470 SG4223 1.614 1.488 2.122 3.121 2.084 1.895 1.033  
> MET508 SD4594 0.992 0.736 2.141 3.650 2.247 1.866 1.463  
> CYS526 SG4769 4.030 3.790 3.437 2.925 3.738 3.605 4.348  
> MET527 SD4777 3.243 3.003 2.748 2.444 3.027 2.875 3.490  
> HIS377 HIS400 HIS410 HIS468 CYS470 MET508 CYS526  
> NE23319 NE23554 NE23646 NE24198 SG4223 SD4594 SG4769  
> HIS400 NE23554 2.048  
> HIS410 NE23646 1.410 2.634  
> HIS468 NE24198 1.862 2.028 1.854  
> CYS470 SG4223 1.693 2.927 1.304 1.268  
> MET508 SD4594 0.891 2.372 1.917 1.673 1.502  
> CYS526 SG4769 4.611 6.612 4.424 5.351 4.122 4.436  
> MET527 SD4777 3.826 5.782 3.577 4.447 3.210 3.636 0.921
> 
> Start terminus LYS-45: NH3+  
> End terminus PHE-528: COO-  
> Opening force field file ./charmm36-feb2021.ff/merged.arn
> 
> * * *
> 
> Program: gmx pdb2gmx, version 2020.1-Ubuntu-2020.1-1  
> Source file: src/gromacs/gmxpreprocess/pdb2gmx.cpp (line 745)
> 
> Fatal error:  
> Atom HN1 in residue LYS 45 was not found in rtp entry LYS with 24 atoms  
> while sorting atoms.
> 
> For a hydrogen, this can be a different protonation state, or it  
> might have had a different number in the PDB file and was rebuilt  
> (it might for instance have been H3, and we only expected H1 & H2).  
> Note that hydrogens might have been added to the entry for the N-terminus.  
> Remove this hydrogen or choose a different protonation state to solve it.  
> Option -ignh will ignore all hydrogens in the input.
> 
> for more information and tips for troubleshooting, please check the GROMACS  
> website at [Common Errors — GROMACS webpage https://www.gromacs.org documentation](http://www.gromacs.org/Documentation/Errors)

> * * *

Then I have used

` -ignh -missing`

to pass the error.

---

<div class="post-metadata">

**Author:** ![jalemkul](https://dub1.discourse-cdn.com/flex017/user_avatar/gromacs.bioexcel.eu/jalemkul/32/18_2.png) [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)\
**Post date:** [September 2, 2021, 2:22pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/8 "2021-09-02T14:22:43Z")

</div>

Never use `-missing`. Your topology is missing an N-terminal proton that leads to your problem. Use `-ignh` to have H atoms rebuilt and appropriately named to solve your issue.

---

<div class="post-metadata">

**Author:** ![serta](https://avatars.discourse-cdn.com/v4/letter/s/65b543/32.png) [@serta](https://gromacs.bioexcel.eu/u/serta)\
**Post date:** [September 2, 2021, 2:43pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/9 "2021-09-02T14:43:26Z")

</div>

the new problem after using only `-ignh`

> going to rename ./charmm36-feb2021.ff/merged.r2b  
> Opening force field file ./charmm36-feb2021.ff/merged.r2b  
> Reading 5fsah\_clean.pdb…  
> WARNING: all CONECT records are ignored  
> Read ‘’, 3972 atoms  
> Analyzing pdb file  
> Splitting chemical chains based on TER records or chain id changing.  
> There are 2 chains and 0 blocks of water and 485 residues with 3972 atoms
> 
> chain #res #atoms  
> 1 ‘A’ 484 3929  
> 2 ’ ’ 1 43
> 
> WARNING: there were 4 atoms with zero occupancy and 34 atoms with  
> occupancy unequal to one (out of 3972 atoms). Check your pdb file.
> 
> Opening force field file ./charmm36-feb2021.ff/atomtypes.atp  
> Reading residue database… (Charmm36-feb2021)  
> Opening force field file ./charmm36-feb2021.ff/merged.rtp  
> Opening force field file ./charmm36-feb2021.ff/merged.hdb  
> Opening force field file ./charmm36-feb2021.ff/merged.n.tdb  
> Opening force field file ./charmm36-feb2021.ff/merged.c.tdb  
> Processing chain 1 ‘A’ (3929 atoms, 484 residues)  
> Analysing hydrogen-bonding network for automated assignment of histidine  
> protonation. 741 donors and 722 acceptors were found.  
> There are 956 hydrogen bonds  
> Will use HISE for residue 101  
> Will use HISE for residue 120  
> Will use HISE for residue 183  
> Will use HISE for residue 242  
> Will use HISE for residue 283  
> Will use HISE for residue 310  
> Will use HISE for residue 322  
> Will use HISE for residue 328  
> Will use HISE for residue 373  
> Will use HISE for residue 377  
> Will use HISE for residue 400  
> Will use HISE for residue 410  
> Will use HISE for residue 468  
> Identified residue LYS45 as a starting terminus.  
> Identified residue PHE528 as a ending terminus.  
> 8 out of 8 lines of specbond.dat converted successfully  
> Special Atom Distance matrix:  
> CYS75 MET86 MET92 HIS101 HIS120 CYS134 MET140  
> SG262 SD357 SD402 NE2471 NE2625 SG728 SD775  
> MET86 SD357 1.393  
> MET92 SD402 1.275 0.978  
> HIS101 NE2471 1.617 2.461 1.626  
> HIS120 NE2625 2.804 1.962 1.549 2.751  
> CYS134 SG728 3.417 3.052 2.266 2.599 1.448  
> MET140 SD775 3.160 3.184 2.267 1.966 2.149 1.036  
> HIS183 NE21148 4.841 5.294 4.811 4.537 4.983 5.035 5.130  
> MET189 SD1186 2.805 3.092 2.521 2.538 2.776 2.995 3.118  
> MET209 SD1344 4.710 4.555 3.805 3.842 2.988 2.323 2.826  
> HIS242 NE21624 3.071 2.596 2.227 3.090 1.650 2.370 2.984  
> MET258 SD1762 4.631 4.160 3.503 3.913 2.295 1.679 2.528  
> HIS283 NE21974 4.741 4.771 3.855 3.411 3.307 1.915 1.797  
> MET292 SD2043 4.112 3.867 3.039 3.084 2.206 0.872 1.365  
> MET306 SD2149 3.288 3.073 2.375 2.733 1.763 1.682 2.211  
> HIS310 NE22177 2.917 2.882 2.181 2.425 1.957 2.017 2.341  
> HIS322 NE22270 2.430 3.303 2.689 2.005 3.528 3.578 3.297  
> HIS328 NE22317 3.806 4.869 4.212 3.091 5.040 4.785 4.312  
> MET372 SD2673 2.230 3.229 2.594 1.720 3.566 3.575 3.190  
> HIS373 NE22684 2.217 3.041 2.399 1.772 3.237 3.305 3.039  
> MET374 SD2691 1.898 2.572 1.684 0.776 2.524 2.353 1.910  
> HIS377 NE22717 1.495 1.796 1.101 1.460 1.974 2.418 2.384  
> HIS400 NE22906 1.577 0.998 0.966 2.124 1.967 2.687 2.624  
> HIS410 NE22984 1.936 2.815 1.967 0.629 2.939 2.706 2.121  
> HIS468 NE23435 2.583 2.544 1.629 1.611 1.693 1.060 0.641  
> CYS470 SG3452 2.816 3.157 2.197 1.549 2.390 1.693 1.286  
> MET508 SD3762 2.306 2.165 1.455 2.015 1.443 1.822 2.111  
> CYS526 SG3910 5.803 6.354 5.672 5.038 5.738 5.351 5.308  
> MET527 SD3917 5.053 5.597 4.857 4.183 4.933 4.477 4.392  
> HIS183 MET189 MET209 HIS242 MET258 HIS283 MET292  
> NE21148 SD1186 SD1344 NE21624 SD1762 NE21974 SD2043  
> MET189 SD1186 2.312  
> MET209 SD1344 3.661 2.624  
> HIS242 NE21624 3.516 1.601 2.401  
> MET258 SD1762 4.654 3.155 1.237 2.313  
> HIS283 NE21974 5.075 3.669 2.069 3.631 2.114  
> MET292 SD2043 4.943 3.191 1.795 2.730 1.294 1.167  
> MET306 SD2149 3.469 1.581 1.510 1.099 1.614 2.566 1.800  
> HIS310 NE22177 3.107 1.043 1.893 1.082 2.173 2.865 2.213  
> HIS322 NE22270 2.673 1.349 3.584 2.796 4.158 4.050 3.813  
> HIS328 NE22317 2.781 2.714 4.412 4.280 5.248 4.665 4.811  
> MET372 SD2673 3.063 1.679 3.795 3.017 4.309 4.056 3.847  
> HIS373 NE22684 2.839 1.231 3.439 2.578 3.942 3.869 3.577  
> MET374 SD2691 3.829 1.813 3.187 2.513 3.402 3.081 2.733  
> HIS377 NE22717 3.730 1.471 3.252 1.776 3.308 3.625 2.966  
> HIS400 NE22906 5.768 3.486 4.580 3.055 4.098 4.359 3.532  
> HIS410 NE22984 3.975 2.124 3.540 2.958 3.803 3.279 3.055  
> HIS468 NE23435 4.969 2.809 2.970 2.601 2.636 2.344 1.693  
> CYS470 SG3452 3.895 2.011 2.356 2.446 2.595 2.064 1.818  
> MET508 SD3762 3.651 1.376 2.475 1.091 2.446 3.090 2.298  
> CYS526 SG3910 1.703 3.287 3.652 4.488 4.854 4.752 5.006  
> MET527 SD3917 1.752 2.598 2.947 3.816 4.118 3.906 4.149  
> MET306 HIS310 HIS322 HIS328 MET372 HIS373 MET374  
> SD2149 NE22177 NE22270 NE22317 SD2673 NE22684 SD2691  
> HIS310 NE22177 0.565  
> HIS322 NE22270 2.624 2.081  
> HIS328 NE22317 3.903 3.426 1.637  
> MET372 SD2673 2.807 2.283 0.413 1.642  
> HIS373 NE22684 2.402 1.863 0.302 1.897 0.455  
> MET374 SD2691 2.089 1.725 1.478 2.720 1.318 1.212  
> HIS377 NE22717 1.836 1.425 1.664 3.251 1.664 1.372 1.050  
> HIS400 NE22906 3.215 3.100 3.546 4.993 3.388 3.265 2.452  
> HIS410 NE22984 2.525 2.153 1.478 2.515 1.209 1.268 0.458  
> HIS468 NE23435 2.024 2.076 3.006 4.205 2.898 2.724 1.584  
> CYS470 SG3452 1.614 1.488 2.122 3.121 2.084 1.895 1.033  
> MET508 SD3762 0.992 0.736 2.141 3.650 2.247 1.866 1.463  
> CYS526 SG3910 4.030 3.790 3.437 2.925 3.738 3.605 4.348  
> MET527 SD3917 3.243 3.003 2.748 2.444 3.027 2.875 3.490  
> HIS377 HIS400 HIS410 HIS468 CYS470 MET508 CYS526  
> NE22717 NE22906 NE22984 NE23435 SG3452 SD3762 SG3910  
> HIS400 NE22906 2.048  
> HIS410 NE22984 1.410 2.634  
> HIS468 NE23435 1.862 2.028 1.854  
> CYS470 SG3452 1.693 2.927 1.304 1.268  
> MET508 SD3762 0.891 2.372 1.917 1.673 1.502  
> CYS526 SG3910 4.611 6.612 4.424 5.351 4.122 4.436  
> MET527 SD3917 3.826 5.782 3.577 4.447 3.210 3.636 0.921
> 
> Start terminus LYS-45: NH3+  
> End terminus PHE-528: COO-  
> Opening force field file ./charmm36-feb2021.ff/merged.arn  
> Checking for duplicate atoms…  
> Generating any missing hydrogen atoms and/or adding termini.  
> Now there are 484 residues with 7799 atoms  
> Chain time…  
> Making bonds…  
> Number of bonds was 7910, now 7910  
> Generating angles, dihedrals and pairs…  
> Before cleaning: 20814 pairs  
> Before cleaning: 21059 dihedrals  
> Keeping all generated dihedrals  
> Making cmap torsions…  
> There are 482 cmap torsion pairs  
> There are 21059 dihedrals, 1297 impropers, 14293 angles  
> 20625 pairs, 7910 bonds and 0 virtual sites  
> Total mass 55608.682 a.m.u.  
> Total charge -1.000 e  
> Writing topology  
> Processing chain 2 (43 atoms, 1 residues)  
> Problem with chain definition, or missing terminal residues.  
> This chain does not appear to contain a recognized chain molecule.  
> If this is incorrect, you can edit residuetypes.dat to modify the behavior.  
> 8 out of 8 lines of specbond.dat converted successfully  
> Special Atom Distance matrix:  
> HEME0 HEME0  
> CAB20 CAC27  
> HEME0 CAC27 0.689  
> HEME0 Fe43 1.007 0.632  
> Opening force field file ./charmm36-feb2021.ff/merged.arn  
> Checking for duplicate atoms…  
> Generating any missing hydrogen atoms and/or adding termini.  
> Now there are 1 residues with 43 atoms  
> Chain time…  
> Making bonds…  
> Warning: Long Bond (1-2 = 0.376383 nm)  
> Warning: Long Bond (1-4 = 0.717554 nm)  
> Warning: Long Bond (1-5 = 0.413299 nm)  
> Warning: Long Bond (2-6 = 0.511743 nm)  
> Warning: Long Bond (6-7 = 0.503469 nm)  
> Warning: Long Bond (7-8 = 0.431017 nm)  
> Warning: Long Bond (8-9 = 0.333521 nm)  
> Warning: Long Bond (7-27 = 0.809707 nm)  
> Warning: Long Bond (27-28 = 0.732382 nm)  
> Warning: Long Bond (28-29 = 0.717554 nm)  
> Warning: Long Bond (29-30 = 0.716079 nm)  
> Warning: Long Bond (29-31 = 0.316329 nm)  
> Warning: Long Bond (8-26 = 0.284035 nm)  
> Warning: Long Bond (23-9 = 0.299013 nm)  
> Warning: Long Bond (23-10 = 0.787275 nm)  
> Warning: Long Bond (3-10 = 0.948874 nm)  
> Warning: Long Bond (10-11 = 1.20847 nm)  
> Warning: Long Bond (11-12 = 1.13427 nm)  
> Warning: Long Bond (12-13 = 0.923594 nm)  
> Warning: Long Bond (3-13 = 0.460306 nm)  
> Warning: Long Bond (11-32 = 1.08176 nm)  
> Warning: Long Bond (12-33 = 0.337877 nm)  
> Warning: Long Bond (33-34 = 0.808382 nm)  
> Warning: Long Bond (24-13 = 0.642844 nm)  
> Warning: Long Bond (24-14 = 0.642262 nm)  
> Warning: Long Bond (4-14 = 0.522709 nm)  
> Warning: Long Bond (14-15 = 0.42877 nm)  
> Warning: Long Bond (15-16 = 0.293668 nm)  
> Warning: Long Bond (4-17 = 0.447375 nm)  
> Warning: Long Bond (15-35 = 0.466005 nm)  
> Warning: Long Bond (25-18 = 0.423339 nm)  
> Warning: Long Bond (18-19 = 0.510099 nm)  
> Warning: Long Bond (20-21 = 0.288529 nm)  
> Warning: Long Bond (19-38 = 0.447402 nm)  
> Warning: Long Bond (20-39 = 0.523604 nm)  
> Warning: Long Bond (39-40 = 0.434792 nm)  
> Warning: Long Bond (41-42 = 0.361641 nm)  
> Warning: Long Bond (41-43 = 0.30647 nm)  
> Warning: Long Bond (22-21 = 0.513304 nm)  
> Warning: Long Bond (22-6 = 0.763283 nm)
> 
> WARNING: atom HA is missing in residue HEME 0 in the pdb file  
> You might need to add atom HA to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HB is missing in residue HEME 0 in the pdb file  
> You might need to add atom HB to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HC is missing in residue HEME 0 in the pdb file  
> You might need to add atom HC to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HD is missing in residue HEME 0 in the pdb file  
> You might need to add atom HD to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMA1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMA1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMA2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMA2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMA3 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMA3 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HAA1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HAA1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HAA2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HAA2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HBA1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HBA1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HBA2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HBA2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMB1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMB1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMB2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMB2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMB3 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMB3 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HAB is missing in residue HEME 0 in the pdb file  
> You might need to add atom HAB to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HBB1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HBB1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HBB2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HBB2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMC1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMC1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMC2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMC2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMC3 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMC3 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HAC is missing in residue HEME 0 in the pdb file  
> You might need to add atom HAC to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HBC1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HBC1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HBC2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HBC2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMD1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMD1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMD2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMD2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HMD3 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HMD3 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HAD1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HAD1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HAD2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HAD2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HBD1 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HBD1 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> WARNING: atom HBD2 is missing in residue HEME 0 in the pdb file  
> You might need to add atom HBD2 to the hydrogen database of building block HEME  
> in the file merged.hdb (see the manual)
> 
> * * *
> 
> Program: gmx pdb2gmx, version 2020.1-Ubuntu-2020.1-1  
> Source file: src/gromacs/gmxpreprocess/pdb2top.cpp (line 1494)
> 
> Fatal error:  
> There were 30 missing atoms in molecule Other, if you want to use this  
> incomplete topology anyhow, use the option -missing
> 
> > For more information and tips for troubleshooting, please check the GROMACS  
> > website at [Common Errors — GROMACS webpage https://www.gromacs.org documentation](http://www.gromacs.org/Documentation/Errors)
> 
> * * *

---

<div class="post-metadata">

**Author:** ![jalemkul](https://dub1.discourse-cdn.com/flex017/user_avatar/gromacs.bioexcel.eu/jalemkul/32/18_2.png) [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)\
**Post date:** [September 2, 2021, 3:03pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/10 "2021-09-02T15:03:01Z")

</div>

Long bonds mean your structure is incomplete or the coordinates are not correct. There is no `.hdb` entry for heme in the C36 port so either you need to fix the H nomenclature and not use `-ignh` or write the `.hdb` entry for heme yourself.

---

<div class="post-metadata">

**Author:** ![serta](https://avatars.discourse-cdn.com/v4/letter/s/65b543/32.png) [@serta](https://gromacs.bioexcel.eu/u/serta)\
**Post date:** [September 2, 2021, 4:52pm UTC](https://gromacs.bioexcel.eu/t/please-how-can-i-neutralize-the-system/2757/11 "2021-09-02T16:52:04Z")

</div>

I have done both, and correct the PDB file, and it worked.  
Thank you very much for you instructions; I appreciate your kind and important assistance.

Best regards.