Prepare PRMTOP and INPCRD for glycoprotein in AMBERTOOLS14.

I used three days to figure out how to do this. If someone could teach me, it would save a lot of time ūüė¶

I will use 4C55 at PDB to introduce this procedure.

  1. Download 4C55 PDB from Protein Data Bank. It contains two chains, some crystal water and a few small molecules.
  2. Use VMD to open it and save it as PDB file. The format will¬†be much easier to edit than the previous one. Select “chain A and not water” to A.pdb, “chain B and not water” to B.pdb.¬†I use A.pdb as example, the process for B.pdb is the same.
  3. Delete the lines of residue EDO and MES in A.pdb, I don’t need them.
  4. sed -i /HOH/a\TER A.pdb Add TER under each line of water oxygen.
  5. Manually add TER at the end of protein chain. In my case, it’s at line 1620, SER444.
  6. ASN297 is the residue to link the glycan with ND2, change the residue name from ASN to NLN. It means the residue links glycan with N.
  7. There are four CYS residues. They will form four S-S bonds. Change the residue name to CYX, so that hydrogen atoms will not be added on S atoms.
  8. The following residues are glycan. Add TER at each glycan residue. They will be linked in tleap later. The residue name in PDB file can not be recognized by tleap according to GLYCAM force field. You need to change it manually. I googled a lot, but I cannot find a table to tell me which is which. I use the carbohydrate builder at¬†http://glycam.org/tools/molecular-dynamics/oligosaccharide-builder/build-glycan?id=1 to do this. I have known NAG is GlcNAc, BMA and MAN are¬†Man, GAL is Gal, FUC is Fuc. That’s enough. I checked the sequence of the glycan can wrote it down. It’s¬†shown below. The number is the number of carbon in the ring. BMA connects three residues with C1, C3, C6. The second NAG at the second line also connects three residues with C1, C4, C6, in which C1 links the protein by ND2 of ASN.#################################################################
    DGalpb1-4DGlcpNAcb1-2DManpb1-6[DGlcpNAcb1-2DManpb1-3]DManpb1-4DGlcpNAcb1-4[LFucpb1-6]DGlcpNAcb1-ND2_ASN
    #######################Glycan Structure in ChainA####################And then use carbohydrate builder to build the same structure, download the optimized structure and see the residue name in GLYCAM_06j-1 (The newest GLYCAM version in AMBERTOOLS14). And then change the residue name according to this structure. Then you need to change the atom name of 4YB, UYB and 0YB (previous NAG in different conditions) to GLYCAM, change C7 to C2N, C8 to CME, O7 to O2N.
  9. OK. The pdb file is prepared well now. And no hydrogen in it. You may use reduce -Trim Protein_H.pdb > Protein_noH.pdb to remove hydrogens. Make sure protein and each glycan residues and water oxygen are divided by TER. Then we will use tleap to link glycan residues, as well as glycan-protein and S-S bond.
  10. The tleap script is below. I use VMD to spend a long time to find the residue number. Please remember VMD starts from 0, AMBERTOOLS start from 1.
    #######################################################

    source leaprc.ff14SB
    source leaprc.GLYCAM_06j-1
    a = loadpdb A.pdb
    savepdb a A_amber.pdb
    #A_amber.pdb contains H atoms and missing heavy atoms.
    g = loadpdb A_amber.pdb
    bond g.25.SG g.85.SG     #S-S bond
    bond g.131.SG g.189.SG #S-S bond
    bond g.61.ND2 g.209.C1 #protein-glycan
    bond g.215.C1 g.214.C2  # the following is connect glycan residues.
    bond g.214.C1 g.211.C3
    bond g.211.C6 g.212.C1
    bond g.212.C2 g.213.C1
    bond g.213.C4 g.216.C1
    bond g.211.C1 g.210.C4
    bond g.210.C1 g.209.C4
    bond g.217.C1 g.209.C6
    addions g Na+ 0
    solvatebox g TIP3PBOX 15.0
    saveamberparm g A.prmtop A.inpcrd
    quit
    

    #######################################################

  11. You may combine chain A and chain B before using tleap or combine them in tleap, either is ok.
  12. Now you have PRMTOP and INPCRD, enjoy your simulation.
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