# Difficulty in generating ION.tpr file using AMBER01 FF and SPC water model **URL:** \ **Category:** User discussions\ **Tags:** forcefield\ **Created:** [July 21, 2023, 7:13am UTC](https://gromacs.bioexcel.eu/t/difficulty-in-generating-ion-tpr-file-using-amber01-ff-and-spc-water-model/6869 "2023-07-21T07:13:27Z")\ **Posts on this page:** 2\ **Page:** 1
**Author:** ![22mspi007](https://avatars.discourse-cdn.com/v4/letter/2/3e96dc/32.png) [@22mspi007](https://gromacs.bioexcel.eu/u/22mspi007)\ **Post date:** [July 21, 2023, 7:13am UTC](https://gromacs.bioexcel.eu/t/difficulty-in-generating-ion-tpr-file-using-amber01-ff-and-spc-water-model/6869/1 "2023-07-21T07:13:27Z")
GROMACS version:2022.5 GROMACS modification: Yes/No Here post your question: During performing the MD Simulation of pdb:4xkx in GROMACS 2022.5. We selected AMBER03 Force Field & SPC Water model It is showing error during generating ION.tpr file ERROR IS: (base) master01@master01-HP-Z640-Workstation:~/Desktop/MD\_GROMACS/MD\_GROMACS\_6$ gmx grompp -f ions.mdp -c box\_sol.gro -p topol.top -o ION.tpr :-) GROMACS - gmx grompp, 2022.5 (-: Executable: /usr/local/gromacs/bin/gmx Data prefix: /usr/local/gromacs Working dir: /home/master01/Desktop/MD\_GROMACS/MD\_GROMACS\_6 Command line: gmx grompp -f ions.mdp -c box\_sol.gro -p topol.top -o ION.tpr NOTE 1 [file ions.mdp]: For a correct single-point energy evaluation with nsteps = 0, use continuation = yes to avoid constraining the input coordinates. Setting the LD random seed to -134223233 WARNING 1 [file LIG.itp, line 9]: Atomtype CB was defined previously (e.g. in the forcefield files), and has now been defined again. This could happen e.g. if you would use a self-contained molecule .itp file that duplicates or replaces the contents of the standard force-field files. You should check the contents of your files and remove such repetition. If you know you should override the previous definition, then you could choose to suppress this warning with -maxwarn. WARNING 2 [file LIG.itp, line 11]: Atomtype F was defined previously (e.g. in the forcefield files), and has now been defined again. This could happen e.g. if you would use a self-contained molecule .itp file that duplicates or replaces the contents of the standard force-field files. You should check the contents of your files and remove such repetition. If you know you should override the previous definition, then you could choose to suppress this warning with -maxwarn. WARNING 3 [file LIG.itp, line 13]: Atomtype CR was defined previously (e.g. in the forcefield files), and has now been defined again. This could happen e.g. if you would use a self-contained molecule .itp file that duplicates or replaces the contents of the standard force-field files. You should check the contents of your files and remove such repetition. If you know you should override the previous definition, then you could choose to suppress this warning with -maxwarn. Generated 2701 of the 2701 non-bonded parameter combinations Generating 1-4 interactions: fudge = 0.5 Generated 2691 of the 2701 1-4 parameter combinations Excluding 3 bonded neighbours molecule type ‘Protein\_chain\_A’ Excluding 3 bonded neighbours molecule type ‘LIG’ Excluding 2 bonded neighbours molecule type ‘SOL’ NOTE 2 [file topol.top, line 53544]: System has non-zero total charge: -7.000000 Total charge should normally be an integer. See [Floating point arithmetic — GROMACS webpage https://www.gromacs.org documentation](http://www.gromacs.org/Documentation/Floating_Point_Arithmetic) for discussion on how close it should be to an integer. NOTE 3 [file topol.top, line 53544]: The bond in molecule-type Protein\_chain\_A between atoms 192 OG and 193 HG has an estimated oscillational period of 9.0e-03 ps, which is less than 10 times the time step of 1.0e-03 ps. Maybe you forgot to change the constraints mdp option. Analysing residue names: There are: 363 Protein residues There are: 1 Other residues There are: 12152 Water residues Analysing Protein… Analysing residues not classified as Protein/DNA/RNA/Water and splitting into groups… Number of degrees of freedom in T-Coupling group rest is 90051.00 NOTE 4 [file ions.mdp]: NVE simulation with an initial temperature of zero: will use a Verlet buffer of 10%. Check your energy drift! NOTE 5 [file ions.mdp]: You are using a plain Coulomb cut-off, which might produce artifacts. You might want to consider using PME electrostatics. This run will generate roughly 3 Mb of data There were 5 notes There were 3 warnings * * * Program: gmx grompp, version 2022.5 Source file: src/gromacs/gmxpreprocess/grompp.cpp (line 2661) Fatal error: Too many warnings (3). If you are sure all warnings are harmless, use the -maxwarn option. ## 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) (base) master01@master01-HP-Z640-Workstation:~/Desktop/MD\_GROMACS/MD\_GROMACS\_6$ gmx grompp -f ions.mdp -c box\_sol.gro -maxwarn 2 -p topol.top -o ION.tpr :-) GROMACS - gmx grompp, 2022.5 (-: Executable: /usr/local/gromacs/bin/gmx Data prefix: /usr/local/gromacs Working dir: /home/master01/Desktop/MD\_GROMACS/MD\_GROMACS\_6 Command line: gmx grompp -f ions.mdp -c box\_sol.gro -maxwarn 2 -p topol.top -o ION.tpr NOTE 1 [file ions.mdp]: For a correct single-point energy evaluation with nsteps = 0, use continuation = yes to avoid constraining the input coordinates. Setting the LD random seed to -214961922 WARNING 1 [file LIG.itp, line 9]: Atomtype CB was defined previously (e.g. in the forcefield files), and has now been defined again. This could happen e.g. if you would use a self-contained molecule .itp file that duplicates or replaces the contents of the standard force-field files. You should check the contents of your files and remove such repetition. If you know you should override the previous definition, then you could choose to suppress this warning with -maxwarn. WARNING 2 [file LIG.itp, line 11]: Atomtype F was defined previously (e.g. in the forcefield files), and has now been defined again. This could happen e.g. if you would use a self-contained molecule .itp file that duplicates or replaces the contents of the standard force-field files. You should check the contents of your files and remove such repetition. If you know you should override the previous definition, then you could choose to suppress this warning with -maxwarn. WARNING 3 [file LIG.itp, line 13]: Atomtype CR was defined previously (e.g. in the forcefield files), and has now been defined again. This could happen e.g. if you would use a self-contained molecule .itp file that duplicates or replaces the contents of the standard force-field files. You should check the contents of your files and remove such repetition. If you know you should override the previous definition, then you could choose to suppress this warning with -maxwarn. Generated 2701 of the 2701 non-bonded parameter combinations Generating 1-4 interactions: fudge = 0.5 Generated 2691 of the 2701 1-4 parameter combinations Excluding 3 bonded neighbours molecule type ‘Protein\_chain\_A’ Excluding 3 bonded neighbours molecule type ‘LIG’ Excluding 2 bonded neighbours molecule type ‘SOL’ NOTE 2 [file topol.top, line 53544]: System has non-zero total charge: -7.000000 Total charge should normally be an integer. NOTE 3 [file topol.top, line 53544]: The bond in molecule-type Protein\_chain\_A between atoms 192 OG and 193 HG has an estimated oscillational period of 9.0e-03 ps, which is less than 10 times the time step of 1.0e-03 ps. Maybe you forgot to change the constraints mdp option. Analysing residue names: There are: 363 Protein residues There are: 1 Other residues There are: 12152 Water residues Analysing Protein… Analysing residues not classified as Protein/DNA/RNA/Water and splitting into groups… Number of degrees of freedom in T-Coupling group rest is 90051.00 NOTE 4 [file ions.mdp]: NVE simulation with an initial temperature of zero: will use a Verlet buffer of 10%. Check your energy drift! NOTE 5 [file ions.mdp]: You are using a plain Coulomb cut-off, which might produce artifacts. You might want to consider using PME electrostatics. This run will generate roughly 3 Mb of data There were 5 notes There were 3 warnings * * * Program: gmx grompp, version 2022.5 Source file: src/gromacs/gmxpreprocess/grompp.cpp (line 2661) Fatal error: Too many warnings (3). If you are sure all warnings are harmless, use the -maxwarn option. * * * ---
**Author:** ![Karis](https://avatars.discourse-cdn.com/v4/letter/k/ed8c4c/32.png) [@Karis](https://gromacs.bioexcel.eu/u/Karis)\ **Post date:** [July 21, 2023, 3:26pm UTC](https://gromacs.bioexcel.eu/t/difficulty-in-generating-ion-tpr-file-using-amber01-ff-and-spc-water-model/6869/2 "2023-07-21T15:26:54Z")
As mentioned in the warnings, there are atoms in lig.itp that were already defined in the force field you’re using. If you want to use the parameters in lig.itp, you can just use the -maxwarn 3 flag as suggested in the warning messages. If you want to use the parameters from the original forcefield, you can remove the lines in the lig.itp file defining those specific atoms (although this would mean the same ligand has some atoms with parameters from the forcefield and some from wherever you got the lig.itp to begin with)