From: Efthymiou, Christos (christos.dereschuk.20_at_ucl.ac.uk)
Date: Tue Jun 28 2022 - 14:26:49 CDT

Hi,

I have run simulations on a protein-protein complex using NAMD and I would like to calculate the free binding energy of the complex. After searching online, one option I found is the CaFE VMD extension. I have followed the instructions to download and install the program and think I have managed to get it to run with some help from this forum, but I am having trouble interpreting the results.

In the manual, it says to run these commands in the terminal (which terminal, I am not sure) and check the output file:

[cid:371375ea-9567-439b-993d-a5d7899a694a]

I have created a script according to the template in the manual:

````
source C:/Users/Christos/vmd.rc
package require cafe 1.0

mmpbsa -top C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/Morikis_Non_PDBFix_20ns_Test4_QwikMD.psf \
-trj J:/pc/Computational_Experiments/Tests/Molecular_Dynamics_Simulations/QwikMD_or_NAMD/2XQW_Starting_Structure/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/MD.dcd \
-out mmpbsa.log \
-par C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/par_all36_carb.prm \
-par C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/par_all36_cgenff.prm \
-par C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/par_all36_lipid.prm \
-par C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/par_all36_na.prm \
-par C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/par_all36_prot.prm \
-par C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/toppar_all36_carb_glycopeptide.str \
-par C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/toppar_water_ions_namd.str \
-com "segname AP1 CP1" \
-rec "segname AP1" \
-lig "segname CP1" \
-first 0 \
-last -1 \
-stride 1000 \
-mm 1 \
-pb 1 \
-pb_exe C:/Users/Christos/Downloads/delphicpp_v8.4.4_windows_x64.exe \
-pb_rad mparse \
-sa 1 \
-sa_rad mparse \
-sa_gamma 0.00542 \
-sa_beta 0.92
quit
````

I run this script in the VMD TK console with source script_name.vmd and the following prints in the console:

````
CaFE) Sanity check
CaFE) Found 5508 atoms for complex
CaFE) Found 4646 atoms for receptor
CaFE) Found 862 atoms for ligand
CaFE) Loaded 10000 frames for complex
CaFE) Generating new trajectory for complex
CaFE) Loaded 10 frames for complex
CaFE) It took 0 days 0 hrs 15 min 52 sec
CaFE) Calculating the MM term
CaFE) It took 0 days 0 hrs 0 min 33 sec
CaFE) Calculating the PB term
Info) ======================
Info) Please cite TopoTools as:
Info) Axel Kohlmeyer & Josh Vermaas, (2019). TopoTools: Release 1.8
Info) https://urldefense.com/v3/__https://doi.org/10.5281/zenodo.598373__;!!DZ3fjg!4B3Uzy_X82ht6_le5xgXx-m1stnIINoqq_-mGWJmIBP3OdvPObwXjPTw7w5ThlcSUcSyKpGs1yrUReICP4slVHkAW-PGWgk$
Info) ======================


````
In the VMD console, this is printed:

````
vmd > {C:\Users\Christos\Downloads\CaFE_Plugin-master\CaFE_Plugin-master\cafe1.0} {C:/Program Files/VMD/scripts/tcl8.6} {C:/Program Files/VMD/scripts} {C:/Program Files/lib} {C:/Program Files/VMD/scripts/tk8.6} {C:/Program Files/VMD/scripts/tk8.6/ttk} {C:/Program Files/VMD/scripts/vmd} {C:/Program Files/VMD/plugins/WIN64/tcl} {C:/Program Files/VMD/plugins/noarch/tcl} {C:/Program Files/VMD/plugins/noarch/tcl/tablelist6.11/scripts} c:/Users/Christos/Documents/vmdStore
psfplugin) Detected a Charmm PSF file
Info) Using plugin psf for structure file C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/Morikis_Non_PDBFix_20ns_Test4_QwikMD.psf
Info) Analyzing structure ...
Info) Atoms: 44191
Info) Bonds: 31315
Info) Angles: 22978 Dihedrals: 14830 Impropers: 957 Cross-terms: 348
Info) Bondtypes: 0 Angletypes: 0 Dihedraltypes: 0 Impropertypes: 0
Info) Residues: 13295
Info) Waters: 12870
Info) Segments: 11
Info) Fragments: 12945 Protein: 2 Nucleic: 0
dcdplugin) detected standard 32-bit DCD file of native endianness
dcdplugin) CHARMM format DCD file (also NAMD 2.1 and later)
Info) Using plugin dcd for coordinates from file J:/pc/Computational_Experiments/Tests/Molecular_Dynamics_Simulations/QwikMD_or_NAMD/2XQW_Starting_Structure/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/MD.dcd
Info) Coordinate I/O rate 10.5 frames/sec, 5 MB/sec, 950.4 sec
Info) Finished with coordinate file J:/pc/Computational_Experiments/Tests/Molecular_Dynamics_Simulations/QwikMD_or_NAMD/2XQW_Starting_Structure/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/MD.dcd.
Info) Opened coordinate file _mmpbsa_com_tmp.dcd for writing.
Info) Finished with coordinate file _mmpbsa_com_tmp.dcd.
psfplugin) Detected a Charmm PSF file
Info) Using plugin psf for structure file C:/Users/Christos/Desktop/Molecular_Dynamics_Setup_Files/CCP19_and_C3d_ChainA_2XQW/Wildtype/Morikis_Non_PDBFix_20ns_Test4/Morikis_Non_PDBFix_20ns_Test4/run/Morikis_Non_PDBFix_20ns_Test4_QwikMD.psf
Info) Analyzing structure ...
Info) Atoms: 44191
Info) Bonds: 31315
Info) Angles: 22978 Dihedrals: 14830 Impropers: 957 Cross-terms: 348
Info) Bondtypes: 0 Angletypes: 0 Dihedraltypes: 0 Impropertypes: 0
Info) Residues: 13295
Info) Waters: 12870
Info) Segments: 11
Info) Fragments: 12945 Protein: 2 Nucleic: 0
dcdplugin) detected standard 32-bit DCD file of native endianness
dcdplugin) CHARMM format DCD file (also NAMD 2.1 and later)
Info) Using plugin dcd for coordinates from file _mmpbsa_com_tmp.dcd
Info) Finished with coordinate file _mmpbsa_com_tmp.dcd.
Info) Opened coordinate file com_mm_tmp.pdb for writing.
Info) Finished with coordinate file com_mm_tmp.pdb.
Info) Opened coordinate file rec_mm_tmp.pdb for writing.
Info) Finished with coordinate file rec_mm_tmp.pdb.
Info) Opened coordinate file lig_mm_tmp.pdb for writing.
Info) Finished with coordinate file lig_mm_tmp.pdb.
````
Checking the files in the working directory, the only new files created are _mmpbsa_com
_tmp.dcd, com_pb_tmp_0.delphi, com_pb_tmp_0.log, and com_pb_tmp_0.pqr. I have checked each file and the com_pb_tmp_0.log file is the only one which seems to mention energy:

````
Energy> Total grid energy : 318767.64 kT
 Energy> Corrected reaction field energy : -8858.08 kT
 Energy> All required energy terms but grid energy : -8858.08 kT
````

That same file also has this error at the bottom:

````
 ......... PROGRAM EXITS SUCCESSFULLY : WITH TOTAL 1 WARNING(S) ........

 ..................... SHOWING 1 OUT OF 1 WARNING(S) ...................
   
[WARNING #1] IN PARAMETER FILE, USER SPECIFIES TO READ PDB FILE IN MODIFIED FORMAT (CHARGE AND SIZE ARE READ FROM THE PDB FILE AS WELL)

````

I am not sure where I can find the free binding energy or if there is something going wrong with the program and that is why I cannot find the binding energy. In the TK console under "Calculating the PB term" it does not give a time taken to do this step, so I am not sure if the PB term is not being calculated. I cannot find any examples online of running this program, so I am having trouble.

I am also open to using other software/programs to find the free binding energy of the protein-protein complex if there is a simpler way to do this, but the CaFE VMD extension is what I have found so far.

I appreciate any troubleshooting advice!


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