# Issues with pdb2gmx for an expanded protein
**URL:** https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517
**Category:** User discussions
**Created:** [July 17, 2020, 10:44am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517 "2020-07-17T10:44:02Z")
**Posts on this page:** 15
**Page:** 1
### Author:  [@wcc6571](https://gromacs.bioexcel.eu/u/wcc6571)
#### Post date: [July 17, 2020, 10:44am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/1 "2020-07-17T10:44:02Z")
GROMACS version: version 2020.1-Ubuntu-2020.1-1
GROMACS modification: Yes/No
Hello,
I expanded 2Y3J four times (pdb given below) on Mercury CCDC. I then ran the following command to create a gro file:
_gmx pdb2gmx -f 2y3j\_exp4.pdb -o 2y3j.gro -water none -ff oplsaa -p topol.top -n index\_pdb2gmx.ndx -ignh_
However, I run into the error:
**Fatal error:**
\*\* Atom OXT in residue MET 6 was not found in rtp entry MET with 17 atoms\*\*
**while sorting atoms.**
I read that this used to be able to be resolved by adding -ignh, however, this command is no longer valid. How can I fix things in this case?
Thank you.
---
### Author:  [@Yogesh](https://gromacs.bioexcel.eu/u/Yogesh)
#### Post date: [July 17, 2020, 10:53am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/2 "2020-07-17T10:53:45Z")
gmx pdb2gmx -f input.pdb -o processed.gro -ignh -ter -water spc
select terminals and forcefield thereafter.
residue names of your protein doesnt match with gromac ff rtp entry. either customize rtp. but that be tiresome. gromacs will keep on throwing errors for each amino acids.
hope it helps.
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### Author:  [@wcc6571](https://gromacs.bioexcel.eu/u/wcc6571)
#### Post date: [July 17, 2020, 10:58am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/3 "2020-07-17T10:58:15Z")
> [@Yogesh](#):
>
> gmx pdb2gmx -f input.pdb -o processed.gro -ignh -ter -water spc
Dear Yogesh,
Thank you for the suggestion. I tried:
gmx pdb2gmx -f 2y3j\_exp4.pdb -o processed.gro -ignh -ter -water tip4p
However, it now gives a dangling bond error:
There is a dangling bond at at least one of the terminal ends. Fix your
coordinate file, add a new terminal database entry (.tdb), or select the
proper existing terminal entry.
Thanks.
---
### Author:  [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)
#### Post date: [July 17, 2020, 11:10am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/4 "2020-07-17T11:10:22Z")
`pdb2gmx` is not detecting the separate chains correctly. Each chain needs either its own unique chain identifier or needs to be separated by `TER`.
---
### Author:  [@wcc6571](https://gromacs.bioexcel.eu/u/wcc6571)
#### Post date: [July 17, 2020, 11:19am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/5 "2020-07-17T11:19:25Z")
Hi Justin,
Each chain is separated by TER, e.g.
**TER 86 MET B 6**
---
### Author:  [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)
#### Post date: [July 17, 2020, 11:32am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/6 "2020-07-17T11:32:39Z")
Please post the entire screen output from `pdb2gmx`.
---
### Author:  [@wcc6571](https://gromacs.bioexcel.eu/u/wcc6571)
#### Post date: [July 17, 2020, 11:34am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/7 "2020-07-17T11:34:02Z")
This is the GROMACS terminal output:
All occupancies are one
Opening force field file /usr/share/gromacs/top/oplsaa.ff/atomtypes.atp
Reading residue database… (Oplsaa)
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.rtp
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.hdb
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.n.tdb
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.c.tdb
Back Off! I just backed up topol.top to ./#topol.top.26#
Processing chain 1 ‘A’ (168 atoms, 24 residues)
Identified residue ALA1 as a starting terminus.
Identified residue MET6 as a ending terminus.
8 out of 8 lines of specbond.dat converted successfully
Special Atom Distance matrix:
MET6 MET6 MET6
SD40 SD82 SD124
MET6 SD82 1.564
MET6 SD124 3.128 1.564
MET6 SD166 4.692 3.128 1.564
Select start terminus type for ALA-1
0: NH3+
1: ZWITTERION\_NH3+ (only use with zwitterions containing exactly one residue)
2: NH2
3: None
0
Start terminus ALA-1: NH3+
Select end terminus type for MET-6
0: COO-
1: ZWITTERION\_COO- (only use with zwitterions containing exactly one residue)
2: COOH
3: None
0
End terminus MET-6: COO-
* * *
Program: gmx pdb2gmx, version 2020.1-Ubuntu-2020.1-1
Source file: src/gromacs/gmxpreprocess/pdb2gmx.cpp (line 745)
Fatal error:
Atom OXT in residue MET 6 was not found in rtp entry MET with 17 atoms
while sorting atoms.
.
---
### Author:  [@Yogesh](https://gromacs.bioexcel.eu/u/Yogesh)
#### Post date: [July 17, 2020, 1:51pm UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/8 "2020-07-17T13:51:03Z")
hi try altering your end terminal residue choice. 0, 0 or 0 2, 2 0, 3 3 etc.
---
### Author:  [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)
#### Post date: [July 17, 2020, 2:53pm UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/9 "2020-07-17T14:53:47Z")
@wcc6571 you left out a lot of screen output about all the WARNINGS of non-sequential residues. The file has a crazy amount of fragmented stuff at the end of it. I suggest you delete redundant and incorrectly formatted coordinate entries.
The actual problem is that you have four chains with the same chain ID. So there are four polypeptides labeled A, each with a Met-6 as the C-terminus (and correctly having OXT). `pdb2gmx` sees residue 6 with another residue following it, which it interprets as meaning that Met-6 is not actually at a terminus:
```
ATOM 40 SD MET A 6 3.159 -0.365 25.912 1.00 52.74 S
ATOM 41 CE MET A 6 1.673 -1.242 26.340 1.00 54.57 C
ATOM 42 OXT MET A 6 3.476 4.532 24.892 1.00 59.23 O
ATOM 43 N ALA A 1 1.807 20.028 44.815 1.00 28.70 N
ATOM 44 CA ALA A 1 2.511 20.987 43.923 1.00 26.55 C
ATOM 45 C ALA A 1 2.923 20.345 42.578 1.00 26.52 C
```
Each hexapeptide needs to be defined as its own chain and needs to be separated by `TER`. If you correct all of those problems, `pdb2gmx` will work.
---
### Author:  [@wcc6571](https://gromacs.bioexcel.eu/u/wcc6571)
#### Post date: [July 18, 2020, 7:54am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/10 "2020-07-18T07:54:59Z")
Thanks for your reply,@jalemkul @Yogesh. I’ve given the four chains different IDs (i.e. A to AF) and added a TER group after each MET 6, in the [PDB file](https://drive.google.com/file/d/1PQKS_upp5BnNM-qbAYgZpwUBzuaMtsQH/view?usp=sharing). I run the same command as before:
_gmx pdb2gmx -f 2y3j\_hbond\_edit3.pdb -o processed.gro -ignh -ter -water tip4p_
Now I get the following error, where GROMACS doesn’t seem to read the different chain IDs:
GROMACS: gmx pdb2gmx, version 2020.1-Ubuntu-2020.1-1
Executable: /usr/bin/gmx
Data prefix: /usr
Working dir: /home/cwc53\_unix/2Y3J\_expanded\_edit2
Command line:
gmx pdb2gmx -f 2y3j\_hbond\_edit3.pdb -o 2Y3J\_processed\_hbondedit3.gro -water tip4p -missing
Select the Force Field:
From ‘/usr/share/gromacs/top’:
1: AMBER03 protein, nucleic AMBER94 (Duan et al., J. Comp. Chem. 24, 1999-2012, 2003)
2: AMBER94 force field (Cornell et al., JACS 117, 5179-5197, 1995)
3: AMBER96 protein, nucleic AMBER94 (Kollman et al., Acc. Chem. Res. 29, 461-469, 1996)
4: AMBER99 protein, nucleic AMBER94 (Wang et al., J. Comp. Chem. 21, 1049-1074, 2000)
5: AMBER99SB protein, nucleic AMBER94 (Hornak et al., Proteins 65, 712-725, 2006)
6: AMBER99SB-ILDN protein, nucleic AMBER94 (Lindorff-Larsen et al., Proteins 78, 1950-58, 2010)
7: AMBERGS force field (Garcia & Sanbonmatsu, PNAS 99, 2782-2787, 2002)
8: CHARMM27 all-atom force field (CHARM22 plus CMAP for proteins)
9: GROMOS96 43a1 force field
10: GROMOS96 43a2 force field (improved alkane dihedrals)
11: GROMOS96 45a3 force field (Schuler JCC 2001 22 1205)
12: GROMOS96 53a5 force field (JCC 2004 vol 25 pag 1656)
13: GROMOS96 53a6 force field (JCC 2004 vol 25 pag 1656)
14: GROMOS96 54a7 force field (Eur. Biophys. J. (2011), 40, 843-856, DOI: 10.1007/s00249-011-0700-9)
15: OPLS-AA/L all-atom force field (2001 aminoacid dihedrals)
15
Using the Oplsaa force field in directory oplsaa.ff
going to rename oplsaa.ff/aminoacids.r2b
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.r2b
Reading 2y3j\_hbond\_edit3.pdb…
Read ‘CSD ENTRY 0001’, 8 atoms
Analyzing pdb file
Splitting chemical chains based on TER records or chain id changing.
WARNING: Chain identifier ‘0’ is used in two non-sequential blocks.
They will be treated as separate chains unless you reorder your file.
There are 2 chains and 0 blocks of water and 3 residues with 8 atoms
chain #res #atoms
1 ‘0’ 1 4
2 ‘0’ 2 4
All occupancy fields zero. This is probably not an X-Ray structure
Opening force field file /usr/share/gromacs/top/oplsaa.ff/atomtypes.atp
Reading residue database… (Oplsaa)
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.rtp
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.hdb
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.n.tdb
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.c.tdb
Back Off! I just backed up topol.top to ./#topol.top.1#
Processing chain 1 ‘0’ (4 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
* * *
Program: gmx pdb2gmx, version 2020.1-Ubuntu-2020.1-1
Source file: src/gromacs/gmxpreprocess/resall.cpp (line 557)
Fatal error:
Residue ‘B20’ not found in residue topology database
For more information and tips for troubleshooting, please check the GROMACS
website at [http://www.gromacs.org/Documentation/Errors](http://www.gromacs.org/Documentation/Errors)
I’m struggling to fix this, would you have any suggestions?
---
### Author:  [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)
#### Post date: [July 19, 2020, 10:48am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/11 "2020-07-19T10:48:57Z")
> [@wcc6571](#):
>
> chain #res #atoms
> 1 ‘0’ 1 4
> 2 ‘0’ 2 4
Your file is still misformatted because instead of A-F, your chain IDs are being read as 0, which are still non-sequential according to the warning.
---
### Author:  [@wcc6571](https://gromacs.bioexcel.eu/u/wcc6571)
#### Post date: [July 19, 2020, 6:07pm UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/12 "2020-07-19T18:07:06Z")
Would you know how to correct for this? Thank you.
---
### Author:  [@Dr\_DBW](https://gromacs.bioexcel.eu/u/Dr_DBW)
#### Post date: [July 19, 2020, 11:26pm UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/13 "2020-07-19T23:26:34Z")
You need to share what the file actually looks like to be able to say that. There is something wrong with the manner your input file is formatted, it is not how pdb2gmx is expecting, so it is not reading the information out “correctly” or how you think it should.
---
### Author:  [@wcc6571](https://gromacs.bioexcel.eu/u/wcc6571)
#### Post date: [July 21, 2020, 11:12am UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/14 "2020-07-21T11:12:57Z")
@Dr_DBW
This is the pdb I’m working with: [https://drive.google.com/file/d/1QYg2GkNbCp2Ja31aBjLMGL07099mOKhE/view?usp=sharing](https://drive.google.com/file/d/1QYg2GkNbCp2Ja31aBjLMGL07099mOKhE/view?usp=sharing)
I’ve attached the command output. I don’t understand why it cannot detect the different protein chains, even though I manually added on a TER group. It’s definitely a formatting issue, because I quadrupled the protein structure on Mercury (CDCC software) beforehand, and things work on the single 2Y3J structure. Would really appreciate it if you could have a look.
Command line:
gmx pdb2gmx -f 2y3j\_exp.pdb -o processed\_exp.gro -ignh -ter
Select the Force Field:
From ‘/usr/share/gromacs/top’:
1: AMBER03 protein, nucleic AMBER94 (Duan et al., J. Comp. Chem. 24, 1999-2012, 2003)
2: AMBER94 force field (Cornell et al., JACS 117, 5179-5197, 1995)
3: AMBER96 protein, nucleic AMBER94 (Kollman et al., Acc. Chem. Res. 29, 461-469, 1996)
4: AMBER99 protein, nucleic AMBER94 (Wang et al., J. Comp. Chem. 21, 1049-1074, 2000)
5: AMBER99SB protein, nucleic AMBER94 (Hornak et al., Proteins 65, 712-725, 2006)
6: AMBER99SB-ILDN protein, nucleic AMBER94 (Lindorff-Larsen et al., Proteins 78, 1950-58, 2010)
7: AMBERGS force field (Garcia & Sanbonmatsu, PNAS 99, 2782-2787, 2002)
8: CHARMM27 all-atom force field (CHARM22 plus CMAP for proteins)
9: GROMOS96 43a1 force field
10: GROMOS96 43a2 force field (improved alkane dihedrals)
11: GROMOS96 45a3 force field (Schuler JCC 2001 22 1205)
12: GROMOS96 53a5 force field (JCC 2004 vol 25 pag 1656)
13: GROMOS96 53a6 force field (JCC 2004 vol 25 pag 1656)
14: GROMOS96 54a7 force field (Eur. Biophys. J. (2011), 40, 843-856, DOI: 10.1007/s00249-011-0700-9)
15: OPLS-AA/L all-atom force field (2001 aminoacid dihedrals)
15
Using the Oplsaa force field in directory oplsaa.ff
Opening force field file /usr/share/gromacs/top/oplsaa.ff/watermodels.dat
Select the Water Model:
1: TIP4P TIP 4-point, recommended
2: TIP4PEW TIP 4-point with Ewald
3: TIP3P TIP 3-point
4: TIP5P TIP 5-point (see [http://redmine.gromacs.org/issues/1348](http://redmine.gromacs.org/issues/1348) for issues)
5: TIP5P TIP 5-point improved for Ewald sums
6: SPC simple point charge
7: SPC/E extended simple point charge
8: None
1
going to rename oplsaa.ff/aminoacids.r2b
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.r2b
Reading 2y3j\_exp.pdb…
Read ‘AMYLOID BETA A4 PROTEIN; 6 AMYLOID PROTEIN, CEREBRAL VASCULAR AMYLOID PEPTIDE, CVAP, PREA4,; 7 PROTEASE NEXIN-II, PN-II’, 8 atoms
Analyzing pdb file
Splitting chemical chains based on TER records or chain id changing.
WARNING: Chain identifier ‘0’ is used in two non-sequential blocks.
They will be treated as separate chains unless you reorder your file.
There are 2 chains and 0 blocks of water and 4 residues with 8 atoms
chain #res #atoms
1 ‘0’ 2 4
2 ‘0’ 2 4
All occupancy fields zero. This is probably not an X-Ray structure
Opening force field file /usr/share/gromacs/top/oplsaa.ff/atomtypes.atp
Reading residue database… (Oplsaa)
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.rtp
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.hdb
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.n.tdb
Opening force field file /usr/share/gromacs/top/oplsaa.ff/aminoacids.c.tdb
Back Off! I just backed up topol.top to ./#topol.top.22#
Processing chain 1 ‘0’ (4 atoms, 2 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
* * *
Program: gmx pdb2gmx, version 2020.1-Ubuntu-2020.1-1
Source file: src/gromacs/gmxpreprocess/resall.cpp (line 557)
Fatal error:
Residue ‘B20’ not found in residue topology database
---
### Author:  [@jalemkul](https://gromacs.bioexcel.eu/u/jalemkul)
#### Post date: [July 21, 2020, 2:33pm UTC](https://gromacs.bioexcel.eu/t/issues-with-pdb2gmx-for-an-expanded-protein/517/15 "2020-07-21T14:33:11Z")
The spacing of the file is totally wrong and you have an unknown residue as the error states. PDB is a fixed-column format so you need to properly align all the fields.