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