calcium metal nomenclature: Rosetta_cm confusing HETATM CA (calcium) with ATOM CA (alpha-carbon)

[Anonymous]

Dear community,

I had this posted under a different sub-forum but I realized it didn’t fit that forum so deleted it and reposting here.

             Hopefully this is a simple answer but I couldn’t find any answer and I don’t remember this happening on a previous rosetta version, just updated from 2019 to 2021.16.61629_bundle. The goal is to add constraints to the amino acids that bind calcium in a homology model since they are in a small loop and can reorient without calcium present. I’m using the metal_setup mover to apply constraints but I’m open to other suggestions.

 

           I’m doing a homology model of a 4-unit symmetrical membrane channel coupled with Calmodulin. The channel and Calmodulin are seperate chains which is denoted in the fasta with a “/” in the sequence to show the split. All this works fine for un-calcified calmodulin. However, now I’m making a calcified calmodulin structure and I have calcium ions in the single unit INPUT.pdb (output of make_symm.sh file. The thread is made with no issue. The problem is the next step of using rosetta_CM and the xml. I’m using the metals setup mover line (1) to denote the amino acids and residues. However, the calcium ions are being confused by roestta for Alpha-Carbons. I have three Calciums per subunit and they are all being overlayed onto the last alpha carbon in the protein sequence for each single unit (2). 

       Is there a special residue or atom name the rosetta is expecting for Calciums? currently the only naming scheme that works is “CA”. (3)

 

Thanks for any help and let me know if you need any more information,

Ryan 

1. calcium is residues 526-528. interacting residues is 397-408 –> 526, 433-433 –> 527, 470-480 –> 528

<ROSETTASCRIPTS>

    <TASKOPERATIONS>

        <ExtraRotamersGeneric name=”extrachi” ex1=”1″ ex2=”1″ />

    </TASKOPERATIONS>

    <SCOREFXNS>

        <ScoreFunction name=”stage1″ weights=”stage1_membrane.wts” symmetric=”1″>

            <Reweight scoretype=”atom_pair_constraint” weight=”1″/>

        </ScoreFunction>

        <ScoreFunction name=”stage2″ weights=”stage2_membrane.wts” symmetric=”1″>

            <Reweight scoretype=”atom_pair_constraint” weight=”0.5″/>

        </ScoreFunction>

        <ScoreFunction name=”fullatom” weights=”stage3_rlx_membrane.wts” symmetric=”1″>

            <Reweight scoretype=”atom_pair_constraint” weight=”0.5″/>

        </ScoreFunction>

        <ScoreFunction name=”r15″ weights=”ref2015″ >

            <Reweight scoretype=”cart_bonded” weight=”0.5″/>

            <Reweight scoretype=”pro_close” weight=”0.0″/>

         </ScoreFunction>

    </SCOREFXNS>

    <FILTERS>

    </FILTERS>

    <MOVERS>

        <SetupMetalsMover name=”setup_metals” metals_detection_LJ_multiplier=”1.0″ contact_resnums=”239-245,397-408,433-443,470-480″ metal_resnums=”526-528″ />

        <Hybridize name=”hybridize” stage1_scorefxn=”stage1″ stage2_scorefxn=”stage2″ fa_scorefxn=”fullatom” batch=”1″ stage1_increase_cycles=”1.0″ stage2_increase_cycles=”1.0″ add_hetatm=”1″ >

            <Fragments three_mers=”hsk2o_03_05.200_v1_3″ nine_mers=”hsk2o_09_05.200_v1_3″/>

            <Template pdb=”hsk2o-thread-cam-chain-ABCD_INPUT.pdb” cst_file=”AUTO” weight=”1.000″ symmdef=”hsk2o-thread-cam-chain-ABCD.symm” />

        </Hybridize>

        <FastRelax name=”relax” scorefxn=”r15″ task_operations=”extrachi” cartesian=”true” bondangle=”true” bondlength=”true”/>

    </MOVERS>

    <PROTOCOLS>

        <Add mover=”setup_metals”/>

        <Add mover=”hybridize”/>

        <Add mover=”relax”/>

    </PROTOCOLS>

    <OUTPUT scorefxn=”r15″/>

</ROSETTASCRIPTS>

2. notice that the Calciums have the same coordinates as the first alpha-carbon of chain B the start of calmodulin. this is true for all 4 symmetrical units

 

ATOM   6185  OD1 ASN A 379      -2.989   8.407  56.731  1.00  0.00           O

ATOM   6186  ND2 ASN A 379      -3.678   8.654  54.615  1.00  0.00           N

ATOM   6187  H   ASN A 379      -5.605   5.733  55.922  1.00  0.00           H

ATOM   6188  HA  ASN A 379      -4.757   7.483  58.184  1.00  0.00           H

ATOM   6189 1HB  ASN A 379      -5.936   8.097  55.462  1.00  0.00           H

ATOM   6190 2HB  ASN A 379      -5.666   9.317  56.708  1.00  0.00           H

ATOM   6191 1HD2 ASN A 379      -2.719   8.727  54.264  1.00  0.00           H

ATOM   6192 2HD2 ASN A 379      -4.442   8.704  53.978  1.00  0.00           H

TER

ATOM   6194  N   ASP B 380      -1.237 -47.247  33.849  1.00  0.00           N

ATOM   6195  CA  ASP B 380      -2.006 -46.260  34.601  1.00  0.00           C

ATOM   6196  C   ASP B 380      -2.779 -45.354  33.619  1.00  0.00           C

ATOM   6197  O   ASP B 380      -3.335 -45.841  32.634  1.00  0.00           O

ATOM   6198  CB  ASP B 380      -1.035 -45.467  35.514  1.00  0.00           C

ATOM   6199  CG  ASP B 380      -1.668 -44.574  36.608  1.00  0.00           C

ATOM   6200  OD1 ASP B 380      -2.103 -44.975  37.659  1.00  0.00           O

…

HETATM 8425 CA    CA C 526      -2.006 -46.260  34.601  1.00  0.00          CA

HETATM 8426 CA    CA C 527      -2.006 -46.260  34.601  1.00  0.00          CA

HETATM 8427 CA    CA C 528      -2.006 -46.260  34.601  1.00  0.00          CA

 

3.  last few lines of input file. notice no TER line although I’ve tried it with. notice different XYZ coordinates for all atoms. I’ve also tried adjusting the spacing of the “CA” in the residue column. Calmodulin for the input as chain A but converted to chain B due to the fasta file input described above.

ATOM   5840  CA  ASP A 380      -5.804 -51.544  37.679  1.00  0.00           C

…

ATOM   8068 3HB  ALA A 525     -34.729 -39.328  36.187  1.00  0.00           H

HETATM 8069  CA   CA A 526       2.182 -32.441  49.215  1.00239.49

HETATM 8070  CA   CA A 527      -8.813 -37.633  50.564  1.00182.89

HETATM 8071  CA   CA A 528     -33.639 -23.142  47.008  1.00223.54

 

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