Dear List,
I am trying to prepare required files for MD simulation of a ligand-enzyme complex, but am having few problems. To fix the problem, I am now just working on the ligand, because I have fixed the problems associated with the receptor. I have extracted the coordinate lines corresponding to the ligand into a file and then applied "reduce" command. My ligand already have got the hydrogens and 0 hydrogens were added (however, it gives lots of WARNINGs) and the required file was saved, which is basically same as the input file. When I remove the hydrogens, the reduce command does not add hydrogens and again copies the input file into the output file. I guess, this is the origin of the problem I am facing with in the next step, as I will mention later.
$ reduce lig.pdb > temp_h.pdb
reduce: version 3.24 07/24/2013, Copyright 1997-2013, J. Michael Word
Processing file: "lig.pdb"
Database of HETATM connections: "/home/siavoush/amber14/dat/reduce_wwPDB_het_dict.txt"
VDW dot density = 16/A^2
Orientation penalty scale = 1 (100%)
Eliminate contacts within 3 bonds.
Ignore atoms with |occupancy| <= 0.01 during adjustments.
Waters ignored if B-Factor >= 40 or |occupancy| < 0.66
Aromatic rings in amino acids accept hydrogen bonds.
Building or keeping OH & SH Hydrogens.
Rotating NH3 Hydrogens.
Processing Met methyls.
WARNING: atom H171 from LIG will be treated as hydrogen
WARNING: atom H171 from LIG will be treated as hydrogen
WARNING: atom H172 from LIG will be treated as hydrogen
WARNING: atom H172 from LIG will be treated as hydrogen
WARNING: atom H171 from LIG will be treated as hydrogen
WARNING: atom H172 from LIG will be treated as hydrogen
WARNING: atom HC10 from LIG will be treated as hydrogen
WARNING: atom HC10 from LIG will be treated as hydrogen
WARNING: atom HC13 from LIG will be treated as hydrogen
WARNING: atom HC13 from LIG will be treated as hydrogen
WARNING: atom HC15 from LIG will be treated as hydrogen
WARNING: atom HC15 from LIG will be treated as hydrogen
WARNING: atom HC22 from LIG will be treated as hydrogen
WARNING: atom HC22 from LIG will be treated as hydrogen
WARNING: atom HC24 from LIG will be treated as hydrogen
WARNING: atom HC24 from LIG will be treated as hydrogen
WARNING: atom HC26 from LIG will be treated as hydrogen
WARNING: atom HC26 from LIG will be treated as hydrogen
WARNING: atom HC28 from LIG will be treated as hydrogen
WARNING: atom HC28 from LIG will be treated as hydrogen
WARNING: atom HC28 from LIG will be treated as hydrogen
WARNING: atom HC28 from LIG will be treated as hydrogen
WARNING: atom HC26 from LIG will be treated as hydrogen
WARNING: atom HC26 from LIG will be treated as hydrogen
WARNING: atom HC24 from LIG will be treated as hydrogen
WARNING: atom HC24 from LIG will be treated as hydrogen
WARNING: atom HC22 from LIG will be treated as hydrogen
WARNING: atom HC22 from LIG will be treated as hydrogen
WARNING: atom HC15 from LIG will be treated as hydrogen
WARNING: atom HC15 from LIG will be treated as hydrogen
WARNING: atom HC13 from LIG will be treated as hydrogen
WARNING: atom HC13 from LIG will be treated as hydrogen
WARNING: atom HC10 from LIG will be treated as hydrogen
WARNING: atom HC10 from LIG will be treated as hydrogen
WARNING: atom H172 from LIG will be treated as hydrogen
WARNING: atom H172 from LIG will be treated as hydrogen
WARNING: atom H171 from LIG will be treated as hydrogen
WARNING: atom H171 from LIG will be treated as hydrogen
WARNING: atom H172 from LIG will be treated as hydrogen
WARNING: atom H171 from LIG will be treated as hydrogen
Found 23 hydrogens (23 hets)
Standardized 0 hydrogens (0 hets)
Added 0 hydrogens (0 hets)
Removed 0 hydrogens (0 hets)
If you publish work which uses reduce, please cite:
Word, et. al. (1999) J. Mol. Biol. 285, 1735-1747.
For more information see
http://kinemage.biochem.duke.edu
Because my ligand structure has the hydrogens, I proceed to the next step using "antechamber" to produce mol2 file.
$ antechamber -i lig_h.pdb -fi pdb -o lig.mol2 -fo mol2 -c bcc -s 2
$ parmchk -i lig.mol2 -f mol2 -o lig.frcmod
Then I do the followings:
tleap -f oldff/leaprc.ff99SB
source leaprc.gaff
> LIG = loadmol2 lig.mol2
Loading Mol2 file: ./lig.mol2
Reading MOLECULE named LIG
> loadamberparams lig.frcmod
Loading parameters: ./lig.frcmod
Reading force field modification type file (frcmod)
Reading title:
remark goes here
> check LIG
Checking 'LIG'....
Checking parameters for unit 'LIG'.
Checking for bond parameters.
Checking for angle parameters.
Unit is OK.
> saveamberparm LIG lig.prmtop lig.rst7
Checking Unit.
Building topology.
Building atom parameters.
Building bond parameters.
Building angle parameters.
Building proper torsion parameters.
Building improper torsion parameters.
total 20 improper torsions applied
Building H-Bond parameters.
Incorporating Non-Bonded adjustments.
Not Marking per-residue atom chain types.
Marking per-residue atom chain types.
(Residues lacking connect0/connect1 -
these don't have chain types marked:
res total affected
LIG 1
)
(no restraints)
> loadamberparams lig.frcmod
Loading parameters: ./lig.frcmod
Reading force field modification type file (frcmod)
Reading title:
remark goes here
> loadoff LIG.lib
Loading library: ./LIG.lib
Because I will use my pdb file of ligand_complex structure, I just load the lig.pdb file and do the rest to make sure when I do my real calculation, it will go OK:
> lig = loadpdb lig.pdb
Loading PDB file: ./lig.pdb
-- residue 584: duplicate [ C] atoms (total 26)
-- residue 584: duplicate [ H] atoms (total 23)
-- residue 584: duplicate [ O] atoms (total 6)
Warning: Atom names in each residue should be unique.
(Same-name atoms are handled by using the first
occurrence and by ignoring the rest.
Frequently duplicate atom names stem from alternate
conformations in the PDB file.)
Added missing heavy atom: .R<LIG 584>.A<O1 7>
Added missing heavy atom: .R<LIG 584>.A<C1 6>
Added missing heavy atom: .R<LIG 584>.A<C2 8>
Added missing heavy atom: .R<LIG 584>.A<C16 24>
Added missing heavy atom: .R<LIG 584>.A<C25 54>
Added missing heavy atom: .R<LIG 584>.A<C3 10>
Added missing heavy atom: .R<LIG 584>.A<C10 17>
Added missing heavy atom: .R<LIG 584>.A<C17 25>
Added missing heavy atom: .R<LIG 584>.A<O2 27>
Added missing heavy atom: .R<LIG 584>.A<C18 26>
Added missing heavy atom: .R<LIG 584>.A<C4 11>
Added missing heavy atom: .R<LIG 584>.A<C11 18>
Added missing heavy atom: .R<LIG 584>.A<C12 19>
Added missing heavy atom: .R<LIG 584>.A<C21 30>
Added missing heavy atom: .R<LIG 584>.A<C19 28>
Added missing heavy atom: .R<LIG 584>.A<C5 12>
Added missing heavy atom: .R<LIG 584>.A<C6 13>
Added missing heavy atom: .R<LIG 584>.A<C13 20>
Added missing heavy atom: .R<LIG 584>.A<C14 21>
Added missing heavy atom: .R<LIG 584>.A<C22 31>
Added missing heavy atom: .R<LIG 584>.A<C20 29>
Added missing heavy atom: .R<LIG 584>.A<C7 14>
Added missing heavy atom: .R<LIG 584>.A<C8 15>
Added missing heavy atom: .R<LIG 584>.A<C15 22>
Added missing heavy atom: .R<LIG 584>.A<C9 16>
Added missing heavy atom: .R<LIG 584>.A<O5 49>
Added missing heavy atom: .R<LIG 584>.A<C24 50>
Added missing heavy atom: .R<LIG 584>.A<O3 33>
Added missing heavy atom: .R<LIG 584>.A<O4 34>
Added missing heavy atom: .R<LIG 584>.A<C23 35>
total atoms in file: 57
Leap added 52 missing atoms according to residue templates:
30 Heavy
22 H / lone pairs
> saveamberparm lig temp.prmtop temp.rst7
Checking Unit.
WARNING: There is a bond of 9.785598 angstroms between:
------- .R<LIG 584>.A<C15 22> and .R<LIG 584>.A<S 32>
WARNING: There is a bond of 7.735279 angstroms between:
------- .R<LIG 584>.A<C 1> and .R<LIG 584>.A<H 2>
WARNING: There is a bond of 8.151633 angstroms between:
------- .R<LIG 584>.A<C 1> and .R<LIG 584>.A<O 5>
-- ignoring the warnings.
Building topology.
Building atom parameters.
Building bond parameters.
Building angle parameters.
Could not find angle parameter: c1 - c2 - o
Could not find angle parameter: c3 - c1 - c2
Building proper torsion parameters.
Building improper torsion parameters.
total 13 improper torsions applied
Building H-Bond parameters.
Incorporating Non-Bonded adjustments.
Parameter file was not saved.
> savepdb lig temp.pdb
Writing pdb file: temp.pdb
>
As you can see, I failed to generate temp.prmtop and temp.rst7 files.
I am attaching the lig.pdb file just in case it is needed.
I will appreciate it if you could let me know what I am doing wrong.
Cheers,
Siavoush
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Received on Tue Sep 15 2020 - 10:00:03 PDT
