I've done a structure refinement of a protein by Rosetta3.8.
However, I'm not sure that what I've done is correct or not.
The protocol that I followed are list below:
First, I put my protein sequence(.fasta file), torsion angle(predicted by talos+), RDC value(experimental data) into the website( http://robetta.bakerlab.org/fragmentsubmit.jsp) to do fragment generation.
Second, I used the command "minirosetta.default.linuxgccrelease @HGDC_broker_cst.options" to run rosetta.
The file"HGDC_broker_cst.options" are shown below:
#make sure all variable names have been replaced with absolute path and that no line begins with a $ or ~s
-native HGDC.pdb # native PDB file (optional)
-fasta HGDC.fasta # protein sequence in fasta format
-frag3 aat000_03_06.200_v1_3 # protein 3-residue fragments file
-frag9 aat000_09_06.200_v1_3 # protein 9-residue fragments file
-increase_cycles 10 # Increase the number of cycles at each stage in AbinitioRelax by this factor
-rg_reweight 0.5 # Reweight contribution of radius of gyration to total score by this scale factor
-rsd_wt_helix 0.5 # Reweight env, pair, and cb scores for helix residues by this factor
-rsd_wt_loop 0.5 # Reweight env, pair, and cb scores for loop residues by this factor
-reinitialize_mover_for_each_job # jd generate fresh copy of its mover before each apply (once per job)
-find_neighbors_3dgrid # Use a 3D lookup table for doing neighbor calculations. For spherical, well-distributed conformations
-nstruct 1000 # how many structures do you want to generate? Usually want to fold at least 1,000.
-silent HGDC_broker_cst.out # full path to silent file output
-silent_struct_type binary # we want binary silent files
-overwrite # overwrite any existing output with the same name you may have generated
-force_minimize # minimize the structure after making a move, even if no restraints given
Finally, I've got a score.fsc file like this:
SCORE: score fa_atr fa_rep fa_sol fa_intra_rep fa_elec pro_close hbond_sr_bb hbond_lr_bb hbond_bb_sc hbond_sc dslf_fa13 rama omega fa_dun p_aa_pp yhh_planarity ref Filter_Stage2_aBefore Filter_Stage2_bQuarter Filter_Stage2_cHalf Filter_Stage2_dEnd co rms maxsub clashes_total clashes_bb time user_tag description
SCORE: -227.165 -772.819 83.957 444.286 2.071 -84.559 0.478 -18.327 -42.232 -18.666 -13.370 0.000 -8.885 17.856 234.566 -30.453 0.395 -21.463 0.000 0.000 0.000 0.000 27.286 19.088 54.000 0.000 0.000 719.000 002 S_002_00000001
SCORE: -247.325 -770.395 78.062 439.372 2.099 -90.080 0.374 -20.420 -38.220 -12.950 -20.084 0.000 -13.605 15.676 235.575 -31.396 0.130 -21.463 0.000 0.000 0.000 0.000 20.812 15.127 68.000 0.000 0.000 687.000 002 S_002_00000002
My questions are :
1. I don't know what the flags -run:reinitialize_mover_for_each_job & -score:find_neighbors_3dgrid & -out:file:silent_struct_type respectively represents for
should I use them for strcture refinement?
or where can I find the meaning of them in detail
(I've found this page (https://www.rosettacommons.org/docs/latest/full-options-list) that describes the meaning of them but it seems not that clear)
2. In my score.fsc file I can't understand what the fa_atr fa_rep ....... stand for?
Besides, most of the tutorials suggest us compare the score term. But how about other terms? Are they as important as each other?
3. If I want to obtain rmsd of each structures compared to the structure which has the lowest energy, what flags should i use.
(just like what csrosetta online server do)
4. Whether I can refine an inexact structure by inputting a pdb file and other constraint but not reconstruct a strcuture by fragment picking?
I'll really appreciate your advice and suggestions.