REMARKS very slow cooling with increase of vdw evaluate ($seed=849302) set seed $seed end {* read in the PSF file and initial structure *} structure @sry_wt.psf @sry_dna.psf @axis_new.psf @axis_600.psf @axis_700.psf end param @TOPPAR:protein.par @TOPPAR:axes.par @TOPPAR:nucleic.par end coor @srywt_ref.pdb collapse assign (resid 4:81 or resid 105:112 or resid 117:124) 25.0 15.0 {*force, rgyr*} scale 1.0 end {* This collapse term minimizes the difference between the radius of gyration *} {* of the current coordinates for the center part of the protein and the *} {* DNA basepairs and a predicted radius of gyration. *} flags exclude * include bonds angles impr end mini powell nstep=20 end {* an initial minimization of the geometric *} {* constraints only. *} {* set the weights for the experimental energy terms *} evaluate ($knoe = 30.0) {* noes *} evaluate ($asym = 0.1) {* slope of NOE potential *} evaluate ($kcdi = 10.0) {* torsion angles *} evaluate ($krama = 0.002) {*database force constant for protein potentials *} evaluate ($knuc = 0.001) {* database force constant for DNA potentials *} @oriented_pairs.inp {* This is a setup file for dna pair restraints from a *} {* database of dna pairs. *} @plane.inp {* This contains the planarity constraints for the DNA basepairs. *} !--------------------------------------------------------------------------- ! Read experimental restraints !set message off echo off end noe reset nres = 5000 ceiling 1000 ! set message off echo off end class all @6_22noe_prot_fin.tbl @6_22noe_prot_fin_new.tbl @6_30n15noe_arg.tbl @7_2metonly.tbl @helix_hbond.tbl @dna_trial1.tbl @endnoe_dnax.tbl @pp_trial.tbl {* NOEs involving the DNA Phosphates? *} @trial_inter3.tbl @6_22nh_inter.tbl @inter_hb.tbl {* Hydrogen bonds between the protein and the DNA, *} {* including phosphates. *} @ends_bdnazzz.tbl averaging all sum potential all square {* soft *} scale all $knoe sqconstant all 1.0 sqexponent all 2 end noe class dnah @hbonds_dna.tbl {* These hbonds are from high-resolution Xray structures *} {* of individual basepairs, so this is a priori data. *} averaging dnah sum potential dnah biharm scale dnah 10.0 sqexponent dnah 2 sqconstant dnah 1.0 end ! set echo on message on end !set message off echo off end carbon phistep=180 psistep=180 nres=300 class all force 0.5 potential harmonic @C13SHIFTS:rcoil_c13.tbl {* random coil shifts, this is a standard file for *} {* using carbon chemical shifts. *} @C13SHIFTS:expected_edited_c13.tbl {* 13C shift database correlating backbone *} {* torsion angles with secondary chemical shifts.*} {* this is also a standard file. *} @c13_shifts.tbl {* experimental carbon shifts for Ca and Cb. *} end restraints dihed reset scale $kcdi nass = 10000 @dihed_final.tbl {* protein phi/psi dihedral angles *} @dihed_dna_final.tbl @chi_final.tbl {* chi1 and chi2 from protein experiments *} end couplings nres 400 potential harmonic class phi degen 1 force 1.0 coefficients 6.98 -1.38 1.72 -60.0 @hnha_xplr.tbl class e degen 1 force 0.0 coefficients 15.3 -6.2 1.5 120.0 {* 15.3cos2(e+120)-6.2cos(e+120)+1.5 *} @dna_pcoup.tbl {* Couplings involving the DNA Phosphate *} end evaluate ($ksani = 0.01) evaluate ($ksani_CH = 1.0*$ksani) evaluate ($ksani_NCO = 0.050*$ksani) evaluate ($ksani_HNC = 0.108*$ksani) evaluate ($ksani_prot = 1.0*$ksani) evaluate ($ksani_dna = 1.0*$ksani) evaluate ($ksani_CH_dna = 1.0*$ksani) sani nres=1000 class JNH force $ksani potential harmonic coeff 0.0 8.6 0.610 @dip_nh.tbl {* uses axis 500 *} class JCH force $ksani_CH potential harmonic coeff 0.0 8.6 0.610 @dip_caha.tbl {* uses axis 500 *} class JNCO force $ksani_NCO potential harmonic coeff 0.0 8.6 0.610 @dip_nc.tbl {* uses axis 500 *} class JHNC force $ksani_HNC potential harmonic coeff 0.0 8.6 0.610 @dip_hnc.tbl {* uses axis 500 *} class JNHp force $ksani_prot potential harmonic coeff 0.0 8.5 0.60 @17dip_protnh.tbl {* uses axis 600 *} class JNHd force $ksani_dna potential harmonic coeff 0.0 8.5 0.60 @dip_dna_nh.tbl {* uses axis 600 *} class JCHd force $ksani_CH_dna potential harmonic coeff 0.0 8.5 0.60 @final_dna_sug.tbl {* uses axis 600 *} @final_dna_aro.tbl {* uses axis 600 *} end xdip nres=5000 class JCHg {* ribose *} type fixd {* fixed distance between atoms. *} scale 1.0 sign on average sum force $ksani potential harmonic coeff 0.0 8.0 0.59 @CAH_gabriel.tbl {* uses axis 700 *} class HHp type vard {* variable distance between atoms. *} scale 10.46847258 sign off average average force $ksani potential square coeff 0.0 8.0 0.59 @wt_HH_dipy.tbl {* uses axis 700 *} class HHn type vard {* variable distance between atoms. *} scale 10.46847258 sign off average sum force 0.0 !$ksani potential square coeff 0.0 8.0 0.59 @HH_dip_negx.tbl {* uses axis 700 *} end evaluate ($rcon = 0.003) parameters nbonds atom nbxmod 4 {* This is set to 4 (bonds), because we are using *} {* internal dynamics (torsion angle refinement). *} wmin = 0.01 {* warning off *} cutnb = 4.5 {* nonbonded cutoff *} tolerance 0.5 repel= 0.9 {* scale factor for vdW radii = 1 ( L-J radii) *} rexp = 2 {* exponents in (r^irex - R0^irex)^rexp *} irex = 2 rcon=$rcon {* actually set the vdW weight *} end end rama nres=10000 @GAUSSIANS:shortrange_gaussians.tbl @GAUSSIANS:new_shortrange_force.tbl {* @QUARTS:2D_quarts_new.tbl *} {* @QUARTS:3D_quarts_new.tbl *} {* @QUARTS:4D_quarts_intra_new.tbl *} {* @QUARTS:forces_torsion_prot_quarts_intra.tbl *} {* use of the gaussian potentials for protein torsion angle correlation *} {* has yielded better structures than the use of quartic potentials *} {* for the same correlations. *} {* The torsion angles that are being correlated are intraresidue; they *} {* include the mainchain torsion angles, phi and psi, and the sidechain *} {* torsion angles, chi1 and others. Inclusion of this potential has an *} {* improving effect on the agreement of the experimental dihedral angles *} {* with those in the resulting structures. *} @GAUSSIANS:all_na_gaussians.tbl @GAUSSIANS:force_nucleic.tbl end set message on echo on end @GAUSSIANS:newshortrange_setup.tbl {* @QUARTS:setup_quarts_torsions_intra_2D3D.tbl *} {* @QUARTS:setup_quarts_torsions_intra_4D.tbl *} @GAUSSIANS:setup_nucleic_new.tbl vector do (refx=x) (all) vector do (refy=y) (all) vector do (refz=z) (all) dynamics internal reset {* dipolar axes are grouped. *} group (resid 500 ) hinge rotate (resid 500) group (resid 600 ) hinge rotate (resid 600) group (resid 700 ) hinge rotate (resid 700) base (name H5T) evaluate ( $res = 101 ) while ($res le 128) loop dna_sugar break (resid $res and name C4') (resid $res and name O4') hinge bendtorsion (resid $res and name C3') (resid $res and name C2') (resid $res and name C4') hinge bendtorsion (resid $res and name C2') (resid $res and name C1') (resid $res and name C3') hinge bendtorsion (resid $res and name C1') (resid $res and name O4') (resid $res and name C2') evaluate ( $res = $res + 1 ) end loop dna_sugar end {* Must exit the dynamics set-up in order to use vector show command to *} {* identify prolines so they can be broken up. *} evaluate ( $res = 1 ) while ($res le 100) loop protein_proline vector show element (resname) (resid $res) eval ($variable =$result) if ($variable = PRO) then dynamics internal reuse=on {* Reuse to keep the assigned breaking from the sugars. *} break (resid $res and name CD) (resid $res and name N) hinge bendtorsion (resid $res and name CA) (resid $res and name CB) (resid $res and name N) hinge bendtorsion (resid $res and name CB) (resid $res and name CG) (resid $res and name CA) hinge bendtorsion (resid $res and name CG) (resid $res and name CD) (resid $res and name CB) end end if evaluate ( $res = $res + 1 ) end loop protein_proline dynamics internal reuse=on evaluate ($res = 1) while ($res le 128) loop group group (resid $res and resname PHE and (name CG or name CD1 or name CD2 or name CE1 or name CE2 or name CZ)) group (resid $res and resname HIS and (name CG or name ND1 or name CD2 or name CE1 or name NE2)) group (resid $res and resname TYR and (name CG or name CD1 or name CD2 or name CE1 or name CE2 or name CZ)) group (resid $res and resname TRP and (name CG or name CD1 or name CD2 or name NE1 or name CE2 or name CE3 or name CZ2 or name CZ3 or name CH2)) group (resid $res and resname CYT and (name n1 or name c6 or name c5 or name c4 or name n3 or name c2)) {* add n4? *} group (resid $res and resname GUA and (name n9 or name c4 or name n3 or name c2 or name n1 or name c6 or name c5 or name n7 or name c8)) {* add n2? *} group (resid $res and resname ADE and (name n9 or name c4 or name n3 or name c2 or name n1 or name c6 or name c5 or name n7 or name c8)) {* add n6? *} group (resid $res and resname THY and (name n1 or name c6 or name c5 or name c4 or name n3 or name c2 or name cm)) {* add cm? *} evaluate ($res = $res +1) end loop group set message on echo on end cloop=false etol=3.0 auto torsion maxe 10000 end evaluate ($cool_steps = 3000) evaluate ($init_t = 3000.01) evaluate ($tol = $init_t/1000) vector do (mass = 100.0) (not (resid 500 or resid 600 or resid 700)) {* all atom masses are set the same so that *} {* motion of each atom is equally likely in *} {* the dynamics. Note that the dynamics is *} {* not a realistic model of the motion of *} {* the system; it is a method of minimizing *} {* a highly complicated function. *} vector do (mass = 100.0) (resid 500 or resid 600 or resid 700) vector do (fbeta = 10.0) (all) {* coupling to heat bath *} coor copy end {* Generate Structures 1 -> 50 *} evaluate ($count =0) while ($count < 3) loop structure evaluate ($count = $count + 1) vector do (x=xcomp) (all) vector do (y=ycomp) (all) vector do (z=zcomp) (all) evaluate ($ini_rad = 0.9) evaluate ($fin_rad = 0.78) evaluate ($ini_con= 0.004) evaluate ($fin_con= 4.0) evaluate ($ini_ang = 0.4) evaluate ($fin_ang = 1.0) evaluate ($ini_imp = 0.1) evaluate ($fin_imp = 1.0) evaluate ($ini_noe = 2.0) evaluate ($fin_noe = 30.0) evaluate ($knoe = $ini_noe) evaluate ($ini_rama = 0.002) evaluate ($fin_rama = 1.0) evaluate ($krama = $ini_rama) evaluate ($ini_nuc = 1.000) evaluate ($fin_nuc = 1.0) evaluate ($knuc = $ini_nuc) evaluate ($kcdi = 10.0) {* torsion angles *} evaluate ($ini_sani = 0.01) evaluate ($fin_sani = 1.0) evaluate ($ksani = $ini_sani) {* value of force constant for dipolar *} {* couplings *} evaluate ($ksani_CH = 1.0*$ksani) evaluate ($ksani_CACO = 0.035*$ksani) evaluate ($ksani_NCO = 0.050*$ksani) evaluate ($ksani_HNC = 0.108*$ksani) evaluate ($ksani_prot = 1.0*$ksani) evaluate ($ksani_dna = 1.0*$ksani) evaluate ($ksani_CH_dna = 1.0*$ksani) sani class JNH force $ksani end sani class JNCO force $ksani_NCO end sani class JCH force $ksani_CH end sani class JHNC force $ksani_HNC end sani class JNHp force $ksani_prot end sani class JNHd force $ksani_dna end sani class JCHd force $ksani_CH_dna end xdip class JCHg force $ksani end xdip class HHp force $ksani end xdip class HHn force 0.0 end flags exclude * include bonds angl impr vdw noe rama carb cdih coup plan sani orie xdip coll end restraints dihed scale $kcdi end noe scale all $knoe end rama {* @QUARTS:forces_torsion_prot_quarts_intra.tbl *} @GAUSSIANS:new_shortrange_force.tbl @GAUSSIANS:force_nucleic.tbl end evaluate ($rcon = 1.0) parameters nbonds atom nbxmod 4 {* This is set to 4 (bonds), because we are using *} {* internal dynamics, which alters torsion angles not *} {* cartesian coordinates *} wmin = 0.01 {* warning off *} cutnb = 100 {* nonbonded cutoff *} tolerance 45 repel= 1.2 {* scale factor for vdW radii = 1 ( L-J radii) *} {* This is larger, because it only applies to CA *} rexp = 2 {* exponents in (r^irex - R0^irex)^rexp *} irex = 2 rcon=$rcon {* actually set the vdW weight *} end end constraints interaction (not name ca) (all) weights * 1 angl 0.4 impr 0.1 vdw 0 elec 0 end interaction (name ca) (name ca) weights * 1 angl 0.4 impr 0.1 vdw 1.0 end {* Due to this set-up, van der Waals forces are turned off *} {* (except those of the CA atoms) for the next dynamics run of *} {* refinement. This allows atoms to pass though one another in *} {* the high temperature phase of annealing. This increases the *} {* conformational space that can be sampled in the high temperature *} {* phase. However, the choice of dynamics type will also affect the *} {* conformational space accessible. *} end vector do (vx = maxwell($init_t)) (all) vector do (vy = maxwell($init_t)) (all) vector do (vz = maxwell($init_t)) (all) dynamics internal itype=pc6 etol=$tol tbath=$init_t response= 5 nstep=5000 timestep= 0.002 endtime=10 cloop=false nprint=10 end parameters nbonds atom nbxmod 4 {* This is set to 4 (bonds), because we are using *} {* internal dynamics, which preserves bonds and *} {* angles and modifies torsion angles. *} wmin = 0.01 {* warning off *} cutnb = 4.5 {* nonbonded cutoff *} tolerance 0.5 repel= 0.9 {* scale factor for vdW radii = 1 ( L-J radii). This *} {* has been reset to a reasonable value and is used *} {* for all atoms. *} rexp = 2 {* exponents in (r^irex - R0^irex)^rexp *} irex = 2 rcon = 1.0 {* actually set the vdW weight *} end end evaluate ($kcdi = 200) restraints dihed scale $kcdi end evaluate ($final_t = 25) { K } evaluate ($tempstep = 12.5) { K } evaluate ($ncycle = ($init_t-$final_t)/$tempstep) evaluate ($nstep = int($cool_steps*4.0/$ncycle)) evaluate ($endtime = $nstep*0.002) evaluate ($bath = $init_t) evaluate ($k_vdw = $ini_con) evaluate ($k_vdwfact = ($fin_con/$ini_con)^(1/$ncycle)) evaluate ($radius= $ini_rad) evaluate ($radfact = ($fin_rad/$ini_rad)^(1/$ncycle)) evaluate ($k_ang = $ini_ang) evaluate ($ang_fac = ($fin_ang/$ini_ang)^(1/$ncycle)) evaluate ($k_imp = $ini_imp) evaluate ($imp_fac = ($fin_imp/$ini_imp)^(1/$ncycle)) evaluate ($noe_fac = ($fin_noe/$ini_noe)^(1/$ncycle)) evaluate ($knoe = $ini_noe) evaluate ($rama_fac = ($fin_rama/$ini_rama)^(1/$ncycle)) evaluate ($krama = $ini_rama) evaluate ($nuc_fac = ($fin_nuc/$ini_nuc)^(1/$ncycle)) evaluate ($knuc = $ini_nuc) evaluate ($sani_fac = ($fin_sani/$ini_sani)^(1/$ncycle)) evaluate ($ksani = $ini_sani) evaluate ($ksani_CH = 1.0*$ksani) evaluate ($ksani_CACO = 0.035*$ksani) evaluate ($ksani_NCO = 0.050*$ksani) evaluate ($ksani_HNC = 0.108*$ksani) evaluate ($ksani_prot = 1.0*$ksani) evaluate ($ksani_dna = 1.0*$ksani) evaluate ($ksani_CH_dna = 1.0*$ksani) vector do (vx = maxwell($bath)) (all) vector do (vy = maxwell($bath)) (all) vector do (vz = maxwell($bath)) (all) evaluate ($i_cool = 0) while ($i_cool < $ncycle) loop cool evaluate ($i_cool=$i_cool+1) evaluate ($bath = $bath - $tempstep) evaluate ($k_vdw=min($fin_con,$k_vdw*$k_vdwfact)) evaluate ($radius=max($fin_rad,$radius*$radfact)) evaluate ($k_ang = $k_ang*$ang_fac) evaluate ($k_imp = $k_imp*$imp_fac) evaluate ($knoe = $knoe*$noe_fac) evaluate ($krama = $krama*$rama_fac) evaluate ($knuc = $knuc*$nuc_fac) evaluate ($ksani = $ksani*$sani_fac) evaluate ($ksani_CH = 1.0*$ksani) evaluate ($ksani_CACO = 0.035*$ksani) evaluate ($ksani_NCO = 0.050*$ksani) evaluate ($ksani_HNC = 0.108*$ksani) evaluate ($ksani_prot = 1.0*$ksani) evaluate ($ksani_dna = 1.0*$ksani) evaluate ($ksani_CH_dna = 1.0*$ksani) constraints interaction (all) (all) weights * 1 angles $k_ang improper $k_imp end end parameter nbonds cutnb=4.5 rcon=$k_vdw nbxmod=4 repel=$radius end end noe scale all $knoe end sani class JNH force $ksani end sani class JNCO force $ksani_NCO end sani class JCH force $ksani_CH end sani class JHNC force $ksani_HNC end sani class JNHp force $ksani_prot end sani class JNHd force $ksani_dna end sani class JCHd force $ksani_CH_dna end xdip class JCHg force $ksani end xdip class HHp force $ksani end xdip class HHn force 0.0 end !$ksani end rama {* @QUARTS:forces_torsion_prot_quarts_intra.tbl *} @GAUSSIANS:new_shortrange_force.tbl @GAUSSIANS:force_nucleic.tbl end dynamics internal itype=pc6 etol=$tol tbath=$bath response= 5 nprint = 10 nstep=$nstep timestep= 0.002 endtime=$endtime cloop=false end end loop cool dynamics internal itype=powell nstep=20000 maxcalls=20000 nprint=10 etol=1e-7 gtol=0.01 depred=0.001 end mini powell nstep=500 end print threshold=0.5 noe evaluate ($rms_noe=$result) evaluate ($violations_noe=$violations) print threshold=0.3 noe evaluate ($rms_noe_03=$result) evaluate ($viol_03=$violations) print threshold=5. cdih evaluate ($rms_cdih=$result) evaluate ($violations_cdih=$violations) print thres=0.05 bonds evaluate ($rms_bonds=$result) print thres=5. angles evaluate ($rms_angles=$result) print thres=5. impropers evaluate ($rms_impropers=$result) couplings print threshold 1.0 class phi end evaluate ($rms_coup = $result) evaluate ($end_viols = $violations) couplings print threshold 1.0 class gly end evaluate ($rms_coup_g = $result) evaluate ($end_viols_g = $violations) couplings print threshold 1.0 class e end evaluate ($rms_coup_e = $result) evaluate ($end_viols_e = $violations) carbon print threshold = 1.0 end evaluate ($rms_ashift = $rmsca) evaluate ($rms_bshift = $rmscb) evaluate ($viol_shift = $violations) sani print threshold=0.0 class JNH end evaluate ($rms_sani_JNH=$result) evaluate ($R_sani_JNH=$result/12.3) evaluate ($viol_sani_JNH=$violations) sani print threshold=0.0 class JNCO end evaluate ($rms_sani_JNCO=$result*0.11068) evaluate ($R_sani_JNCO=$result/12.3) evaluate ($viol_sani_JNCO=$violations) sani print threshold=0.0 class JCH end evaluate ($rms_sani_JCH=$result*2.08953) evaluate ($R_sani_JCH=$result/12.3) evaluate ($viol_sani_JCH=$violations) sani print threshold=0.0 class JHNC end evaluate ($rms_sani_JHNC=$result*0.329) evaluate ($R_sani_JHNC=$result/12.3) evaluate ($viol_sani_JHNC=$violations) sani print threshold=0.0 class JNHp end evaluate ($rms_sani_JNH_prot=$result) evaluate ($R_sani_JNH_prot=$result/12.11) evaluate ($viol_sani_JNH_prot=$violations) sani print threshold=0.0 class JNHd end evaluate ($rms_sani_JNH_dna=$result) evaluate ($R_sani_JNH_dna=$result/12.11) evaluate ($viol_sani_JNH_dna=$violations) sani print threshold=0.0 class JCHd end evaluate ($rms_sani_JCH_dna=$result*2.08953) evaluate ($R_sani_JCH_dna=$result/12.11) evaluate ($viol_sani_JCH_dna=$violations) xdip print threshold=0.0 class JCHg end evaluate ($rms_xdip_JCHg=$result*2.08953) evaluate ($R_xdip_JCHg=$result/10.86) evaluate ($viol_xdip_JCHg=$violations) xdip print threshold=0.0 class HHp end evaluate ($rms_xdip_HHp=$result) evaluate ($viol_xdip_HHp=$violations) xdip print threshold=0.0 class HHn end evaluate ($rms_xdip_HHn=$result) evaluate ($viol_xdip_HHn=$violations) remarks =============================================================== remarks overall,bonds,angles,improper,vdw,cdih,noe,coup, shift, rama, plane, orient, sani, xdip remarks energies: $ener, $bond, $angl, $impr, $vdw, $cdih, $noe, $coup, $carb, $plan, $orie, remarks $sani, $xdip remarks =============================================================== remarks bonds,angles,impropers,cdih,noe,coup remarks bonds etc: $rms_bonds,$rms_angles,$rms_impropers,$rms_cdih,$rms_noe,$rms_coup remarks shifts RMS a, b: $rms_ashift, $rms_bshift remarks =============================================================== remarks cdih noe noe03A HNHAcoup DNAcoup remarks violations : $violations_cdih $violations_noe $viol_03 $end_viols $end_viols_e remarks shifts: $viol_shift remarks =============================================================== remarks jcoup stats: end_rms end_rms_e remarks rms-d: $rms_coup $rms_coup_e remarks =============================================================== remarks sani NH CH NCO HNCO remarks RMS sani: $rms_sani_JNH $rms_sani_JCH $rms_sani_JNCO $rms_sani_JHNC remarks R factor: $R_sani_JNH $R_sani_JCH $R_sani_JNCO $R_sani_JHNC remarks viol sani: $viol_sani_JNH $viol_sani_JCH $viol_sani_JNCO $viol_sani_JHNC remarks sani NH_prot NH_dna CH_dna remarks RMS sani DNA: $rms_sani_JNH_prot $rms_sani_JNH_dna $rms_sani_JCH_dna remarks R factor DNA: $R_sani_JNH_prot $R_sani_JNH_dna $R_sani_JCH_dna remarks viol sani DNA: $viol_sani_JNH_prot $viol_sani_JNH_dna $viol_sani_JCH_dna remarks xdip CH HHp HHn remarks RMS xdip: $rms_xdip_JCHg $rms_xdip_HHp $rms_xdip_HHn remarks R factor xdip: $R_xdip_JCHg remarks viol xdip: $viol_xdip_JCHg $viol_xdip_HHp $viol_xdip_HHn remarks =============================================================== {====>} {*Name(s) of the family of final structures.*} evaluate ($file = "final_" + encode($count) + ".sa") write coor output= $file end end loop structure stop