Minimize protein-covalent ligand complex with AMBER

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This is for ligands that modify a specific residue.

(1) Make a directory:

mkdir coval_min
cd coval_min/


(2) Download pdb file from the web:

 wget https://files.rcsb.org/view/5YY1.pdb


(3) Manually create 3 files using your favorate text editor: a covalent ligand, a co-facotor and a receptor file.

(3.1) for the receptor, rec.pdb:

Change the covalent cysteine to reduce name CYB.

Here is a modified cystiene residue (this is without a charge and without a hydrogen):

cat CYB.prep 
    0    0    2

CYSTEINE without h and without charge for covalent 
                                                                
 CYB  INT     0                                                 
 CORR OMIT DU   BEG                                             
   0.00000                                                      
   1  DUMM  DU    M    0  -1  -2     0.000     0.000     0.000   0.00000
   2  DUMM  DU    M    1   0  -1     1.449     0.000     0.000   0.00000
   3  DUMM  DU    M    2   1   0     1.522   111.100     0.000   0.00000
   4  N     N     M    3   2   1     1.335   116.600   180.000  -0.41570
   5  H     H     E    4   3   2     1.010   119.800     0.000   0.27190
   6  CA    CX    M    4   3   2     1.449   121.900   180.000   0.02130
   7  HA    H1    E    6   4   3     1.090   109.500   300.000   0.11240
   8  CB    2C    3    6   4   3     1.525   111.100    60.000  -0.12310
   9  HB2   H1    E    8   6   4     1.090   109.500   300.000   0.11120
  10  HB3   H1    E    8   6   4     1.090   109.500    60.000   0.11120
  11  SG    SH    E    8   6   4     1.810   116.000   180.000  -0.23580
  12  C     C     M    6   4   3     1.522   111.100   180.000   0.59730
  13  O     O     E   12   6   4     1.229   120.500     0.000  -0.56790

IMPROPER                                                        
 -M   CA   N    H                                               
 CA   +M   C    O                                               
                                                                
DONE                                                            
STOP

Here is a frcmod file with need parameters:

cat thioe.frcmod 
Nir and trent got the parms from gaff and change the names, for thio-ether
MASS

BOND
c3-SH  225.8    1.8210       SOURCE1     358   0.0075
C -SH  225.8    1.8210       SOURCE1     358   0.0075

ANGLE
2C-SH-c3   60.63       99.92   SOURCE3           14    2.0723
CT-SH-c3   60.63       99.92   SOURCE3           14    2.0723
c3-c3-SH   61.10      112.69   SOURCE3           24    2.1842
hc-c3-SH   42.51      108.76   SOURCE2            3    1.6891
SH-c3-h1   42.51      108.76   SOURCE2            3    1.6891

DIHE
X -c3-SH-X    3    1.000         0.000           3.000      JCC,7,(1986),230

IMPROPER

NONBON

These parameter values were taken from the gaff force field (/nfs/soft/amber/amber14/dat/leap/parm/gaff.dat).

(3.2) for a covalent ligand, lig.pdb:

Keep the side chain of the modified cysteine:

cat 94F_bkup.pdb 
ATOM     89  CA  CYS A  12      29.215  -1.013  17.747  1.00 23.10           C  
ATOM     92  CB  CYS A  12      30.264  -1.172  18.849  1.00 23.68           C  
ATOM     93  SG  CYS A  12      29.632  -1.020  20.503  1.00 25.01           S  
ATOM   1385  C31 94F A 203      16.511  -4.125  24.817  1.00 32.17           C  
ATOM   1386  C30 94F A 203      15.606  -4.260  23.787  1.00 32.36           C  
ATOM   1387  C32 94F A 203      17.840  -3.910  24.531  1.00 31.54           C  
ATOM   1388  C29 94F A 203      16.037  -4.183  22.484  1.00 32.47           C  
ATOM   1389  C19 94F A 203      21.614  -2.199  22.881  1.00 29.12           C  
ATOM   1390  C5  94F A 203      23.964  -5.464  22.540  1.00 29.57           C  
ATOM   1391  C18 94F A 203      22.453  -3.293  22.771  1.00 29.13           C  
ATOM   1392  C23 94F A 203      18.284  -3.829  23.223  1.00 31.43           C  
ATOM   1393  C22 94F A 203      19.692  -3.612  23.019  1.00 29.79           C  
ATOM   1394  C24 94F A 203      17.372  -3.972  22.203  1.00 32.05           C  
ATOM   1395  C3  94F A 203      21.900  -4.565  22.770  1.00 29.39           C  
ATOM   1396  C2  94F A 203      20.533  -4.693  22.895  1.00 29.45           C  
ATOM   1397  C20 94F A 203      20.247  -2.347  23.012  1.00 29.51           C  
ATOM   1398  C7  94F A 203      23.836  -3.189  22.644  1.00 28.94           C  
ATOM   1399  C14 94F A 203      29.536   0.583  20.690  1.00 25.47           C  
ATOM   1400  C13 94F A 203      28.376   1.143  21.474  1.00 26.40           C  
ATOM   1401  C12 94F A 203      27.093   0.850  20.795  1.00 26.10           C  
ATOM   1402  C9  94F A 203      23.979  -0.953  21.731  1.00 28.62           C  
ATOM   1403  C17 94F A 203      25.969  -2.149  22.595  1.00 29.17           C  
ATOM   1404  C10 94F A 203      25.024  -0.390  20.794  1.00 28.12           C  
ATOM   1405  C16 94F A 203      26.678  -0.809  22.586  1.00 28.22           C  
ATOM   1406  C25 94F A 203      17.826  -3.897  20.794  1.00 32.91           C  
ATOM   1407  N4  94F A 203      22.652  -5.665  22.653  1.00 29.29           N  
ATOM   1408  N6  94F A 203      24.595  -4.289  22.534  1.00 29.70           N  
ATOM   1409  N8  94F A 203      24.523  -1.948  22.642  1.00 29.33           N  
ATOM   1410  N11 94F A 203      26.238   0.005  21.476  1.00 27.18           N  
ATOM   1411  O15 94F A 203      26.847   1.362  19.713  1.00 25.34           O  
ATOM   1412  F1  94F A 203      19.995  -5.935  22.878  1.00 29.69           F  
ATOM   1413  F26 94F A 203      16.791  -3.770  19.931  1.00 34.77           F  
ATOM   1414  F27 94F A 203      18.674  -2.862  20.538  1.00 31.92           F  
ATOM   1415  F28 94F A 203      18.495  -5.026  20.445  1.00 33.49           F  
ATOM   1416 CL   94F A 203      19.224  -0.960  23.165  1.00 29.22          CL  

Change and make the numbering and naming consistent:

cat 94F.pdb 
ATOM   1382  CA  LIG A 203      29.215  -1.013  17.747  1.00 23.10           C  
ATOM   1383  CB  LIG A 203      30.264  -1.172  18.849  1.00 23.68           C  
ATOM   1384  SG  LIG A 203      29.632  -1.020  20.503  1.00 25.01           S  
ATOM   1385  C31 LIG A 203      16.511  -4.125  24.817  1.00 32.17           C  
ATOM   1386  C30 LIG A 203      15.606  -4.260  23.787  1.00 32.36           C  
ATOM   1387  C32 LIG A 203      17.840  -3.910  24.531  1.00 31.54           C  
ATOM   1388  C29 LIG A 203      16.037  -4.183  22.484  1.00 32.47           C  
ATOM   1389  C19 LIG A 203      21.614  -2.199  22.881  1.00 29.12           C  
ATOM   1390  C5  LIG A 203      23.964  -5.464  22.540  1.00 29.57           C  
ATOM   1391  C18 LIG A 203      22.453  -3.293  22.771  1.00 29.13           C  
ATOM   1392  C23 LIG A 203      18.284  -3.829  23.223  1.00 31.43           C  
ATOM   1393  C22 LIG A 203      19.692  -3.612  23.019  1.00 29.79           C  
ATOM   1394  C24 LIG A 203      17.372  -3.972  22.203  1.00 32.05           C  
ATOM   1395  C3  LIG A 203      21.900  -4.565  22.770  1.00 29.39           C  
ATOM   1396  C2  LIG A 203      20.533  -4.693  22.895  1.00 29.45           C  
ATOM   1397  C20 LIG A 203      20.247  -2.347  23.012  1.00 29.51           C  
ATOM   1398  C7  LIG A 203      23.836  -3.189  22.644  1.00 28.94           C  
ATOM   1399  C14 LIG A 203      29.536   0.583  20.690  1.00 25.47           C  
ATOM   1400  C13 LIG A 203      28.376   1.143  21.474  1.00 26.40           C  
ATOM   1401  C12 LIG A 203      27.093   0.850  20.795  1.00 26.10           C  
ATOM   1402  C9  LIG A 203      23.979  -0.953  21.731  1.00 28.62           C  
ATOM   1403  C17 LIG A 203      25.969  -2.149  22.595  1.00 29.17           C  
ATOM   1404  C10 LIG A 203      25.024  -0.390  20.794  1.00 28.12           C  
ATOM   1405  C16 LIG A 203      26.678  -0.809  22.586  1.00 28.22           C  
ATOM   1406  C25 LIG A 203      17.826  -3.897  20.794  1.00 32.91           C  
ATOM   1407  N4  LIG A 203      22.652  -5.665  22.653  1.00 29.29           N  
ATOM   1408  N6  LIG A 203      24.595  -4.289  22.534  1.00 29.70           N  
ATOM   1409  N8  LIG A 203      24.523  -1.948  22.642  1.00 29.33           N  
ATOM   1410  N11 LIG A 203      26.238   0.005  21.476  1.00 27.18           N  
ATOM   1411  O15 LIG A 203      26.847   1.362  19.713  1.00 25.34           O  
ATOM   1412  F1  LIG A 203      19.995  -5.935  22.878  1.00 29.69           F  
ATOM   1413  F26 LIG A 203      16.791  -3.770  19.931  1.00 34.77           F  
ATOM   1414  F27 LIG A 203      18.674  -2.862  20.538  1.00 31.92           F  
ATOM   1415  F28 LIG A 203      18.495  -5.026  20.445  1.00 33.49           F  
ATOM   1416 CL   LIG A 203      19.224  -0.960  23.165  1.00 29.22          CL  

Uses chimera to add hydrogens:

Use chimera to add hydrogens


Save the protonated molecule in pdb format and rename the atoms:

cat 94F.full_mod_num.pdb 
HETATM    1  C01 LIG A   1      29.215  -1.013  17.747  1.00  0.00           C
HETATM    2  C02 LIG A   1      30.264  -1.172  18.849  1.00  0.00           C
HETATM    3  S03 LIG A   1      29.632  -1.020  20.503  1.00  0.00           S
HETATM    4  C04 LIG A   1      16.511  -4.125  24.817  1.00  0.00           C
HETATM    5  C05 LIG A   1      15.606  -4.260  23.787  1.00  0.00           C
HETATM    6  C06 LIG A   1      17.840  -3.910  24.531  1.00  0.00           C
HETATM    7  C07 LIG A   1      16.037  -4.183  22.484  1.00  0.00           C
HETATM    8  C08 LIG A   1      21.614  -2.199  22.881  1.00  0.00           C
HETATM    9  C09 LIG A   1      23.964  -5.464  22.540  1.00  0.00           C
HETATM   10  C10 LIG A   1      22.453  -3.293  22.771  1.00  0.00           C
HETATM   11  C11 LIG A   1      18.284  -3.829  23.223  1.00  0.00           C
HETATM   12  C12 LIG A   1      19.692  -3.612  23.019  1.00  0.00           C
HETATM   13  C13 LIG A   1      17.372  -3.972  22.203  1.00  0.00           C
HETATM   14  C14 LIG A   1      21.900  -4.565  22.770  1.00  0.00           C
HETATM   15  C15 LIG A   1      20.533  -4.693  22.895  1.00  0.00           C
HETATM   16  C16 LIG A   1      20.247  -2.347  23.012  1.00  0.00           C
HETATM   17  C17 LIG A   1      23.836  -3.189  22.644  1.00  0.00           C
HETATM   18  C18 LIG A   1      29.536   0.583  20.690  1.00  0.00           C
HETATM   19  C19 LIG A   1      28.376   1.143  21.474  1.00  0.00           C
HETATM   20  C20 LIG A   1      27.093   0.850  20.795  1.00  0.00           C
HETATM   21  C21 LIG A   1      23.979  -0.953  21.731  1.00  0.00           C
HETATM   22  C22 LIG A   1      25.969  -2.149  22.595  1.00  0.00           C
HETATM   23  C23 LIG A   1      25.024  -0.390  20.794  1.00  0.00           C
HETATM   24  C24 LIG A   1      26.678  -0.809  22.586  1.00  0.00           C
HETATM   25  C25 LIG A   1      17.826  -3.897  20.794  1.00  0.00           C
HETATM   26  N26 LIG A   1      22.652  -5.665  22.653  1.00  0.00           N
HETATM   27  N27 LIG A   1      24.595  -4.289  22.534  1.00  0.00           N
HETATM   28  N28 LIG A   1      24.523  -1.948  22.642  1.00  0.00           N
HETATM   29  N29 LIG A   1      26.238   0.005  21.476  1.00  0.00           N
HETATM   30  O30 LIG A   1      26.847   1.362  19.713  1.00  0.00           O
HETATM   31  F31 LIG A   1      19.995  -5.935  22.878  1.00  0.00           F
HETATM   32  F32 LIG A   1      16.791  -3.770  19.931  1.00  0.00           F
HETATM   33  F33 LIG A   1      18.674  -2.862  20.538  1.00  0.00           F
HETATM   34  F34 LIG A   1      18.495  -5.026  20.445  1.00  0.00           F
HETATM   35 CL   LIG A   1      19.224  -0.960  23.165  1.00  0.00          Cl
HETATM   36  H36 LIG A   1      31.026  -0.406  18.705  1.00  0.00           H
HETATM   37  H37 LIG A   1      30.732  -2.151  18.745  1.00  0.00           H
HETATM   38  H38 LIG A   1      16.180  -4.188  25.843  1.00  0.00           H
HETATM   39  H39 LIG A   1      14.561  -4.426  24.003  1.00  0.00           H
HETATM   40  H40 LIG A   1      18.547  -3.803  25.340  1.00  0.00           H
HETATM   41  H41 LIG A   1      15.328  -4.288  21.676  1.00  0.00           H
HETATM   42  H42 LIG A   1      22.038  -1.206  22.864  1.00  0.00           H
HETATM   43  H43 LIG A   1      24.581  -6.345  22.443  1.00  0.00           H
HETATM   44  H44 LIG A   1      29.494   1.022  19.693  1.00  0.00           H
HETATM   45  H45 LIG A   1      30.457   0.921  21.166  1.00  0.00           H
HETATM   46  H46 LIG A   1      28.366   0.694  22.467  1.00  0.00           H
HETATM   47  H47 LIG A   1      28.494   2.222  21.569  1.00  0.00           H
HETATM   48  H48 LIG A   1      23.559  -0.136  22.317  1.00  0.00           H
HETATM   49  H49 LIG A   1      23.184  -1.410  21.141  1.00  0.00           H
HETATM   50  H50 LIG A   1      26.283  -2.718  23.470  1.00  0.00           H
HETATM   51  H51 LIG A   1      26.228  -2.702  21.692  1.00  0.00           H
HETATM   52  H52 LIG A   1      25.273  -1.150  20.054  1.00  0.00           H
HETATM   53  H53 LIG A   1      24.608   0.478  20.282  1.00  0.00           H
HETATM   54  H54 LIG A   1      27.752  -0.975  22.502  1.00  0.00           H
HETATM   55  H55 LIG A   1      26.468  -0.286  23.519  1.00  0.00           H
HETATM   56  H56 LIG A   1      29.692  -1.121  16.773  1.00  0.00           H
HETATM   57  H57 LIG A   1      28.758  -0.026  17.820  1.00  0.00           H
HETATM   58  H58 LIG A   1      28.448  -1.778  17.863  1.00  0.00           H
CONECT   23   21   29   52   53
CONECT   20   19   29   30
CONECT   19   18   20   46   47
CONECT   18    3   19   44   45
CONECT   24   22   29   54   55
CONECT   22   24   28   50   51
CONECT   10    8   14   17
CONECT    8   10   16   42
CONECT   15   12   14   31
CONECT   16    8   12   35
CONECT   12   11   15   16
CONECT   11    6   12   13
CONECT   13    7   11   25
CONECT   25   13   32   33   34
CONECT    7    5   13   41
CONECT   14   10   15   26
CONECT    5    7    4   39
CONECT    4    6    5   38
CONECT    6    4   11   40
CONECT    9   26   27   43
CONECT   17   10   27   28
CONECT   21   23   28   48   49
CONECT    1    2   57   56   58
CONECT    2    1    3   36   37
CONECT   35   16
CONECT   31   15
CONECT   32   25
CONECT   33   25
CONECT   34   25
CONECT   52   23
CONECT   53   23
CONECT   46   19
CONECT   47   19
CONECT   44   18
CONECT   45   18
CONECT   54   24
CONECT   55   24
CONECT   50   22
CONECT   51   22
CONECT   42    8
CONECT   41    7
CONECT   39    5
CONECT   38    4
CONECT   40    6
CONECT   43    9
CONECT   48   21
CONECT   49   21
CONECT   56    1
CONECT   57    1
CONECT   58    1
CONECT   36    2
CONECT   37    2
CONECT   29   20   23   24
CONECT   26    9   14
CONECT   27    9   17
CONECT   28   17   21   22
CONECT   30   20
CONECT    3    2   18
END

Run the following script:

cat run.002.ligprep.antechamber.csh 
#! /bin/tcsh


set workdir = `pwd`
cd $workdir


setenv AMBERHOME /nfs/soft/amber/amber14

rm lig; mkdir lig; cd lig

cp $workdir/94F.full_mod_num.pdb lig.pdb

$AMBERHOME/bin/antechamber -i lig.pdb -fi pdb -o lig.ante.mol2 -fo mol2 

$AMBERHOME/bin/antechamber -i lig.ante.mol2 -fi mol2 -o lig.ante.charge.mol2 -fo mol2 -c bcc -at sybyl -nc 0
$AMBERHOME/bin/antechamber -i lig.ante.mol2 -fi mol2  -o lig.ante.pdb  -fo pdb
$AMBERHOME/bin/antechamber -i lig.ante.charge.mol2 -fi mol2  -o lig.ante.charge.prep -fo prepi
$AMBERHOME/bin/parmchk -i lig.ante.charge.prep -f  prepi -o lig.ante.charge.frcmod


Modify the prep file to remove the cysteine side chain. Also add partial charge to covalent carbon to make the Cysteine (CYB) + covalent ligand an integer.

diff lig/lig.ante.charge.prep lig/lig.ante.charge.mod.prep
11,19c11
<    4  C01   c3    M    3   2   1     1.540   111.208  -180.000 -0.104100
<    5  H56   hc    E    4   3   2     1.090   115.954    83.673  0.047367
<    6  H57   hc    E    4   3   2     1.090    65.214  -175.770  0.047367
<    7  H58   hc    E    4   3   2     1.089    45.251    -8.622  0.047367
<    8  C02   c3    M    4   3   2     1.530   133.294   -81.790 -0.002300
<    9  H36   h1    E    8   4   3     1.090   108.130  -134.600  0.074200
<   10  H37   h1    E    8   4   3     1.090   108.604   107.772  0.074200
<   11  S03   ss    M    8   4   3     1.777   114.690   -13.854 -0.332200
<   12  C18   c3    M   11   8   4     1.617   102.332   -80.293 -0.008300
---
>   12  C18   c3    M    3   2   1     1.617   102.332   -80.293 -0.037200 

(3.3) for a co-factory (gdp), gdp.pdb:

Add hydrogens to gdp is chimera.

Build the prep and frcmod files with antechamber:

cat run.002.ligprep.antechamber_gdp.csh 
#! /bin/tcsh


set workdir = `pwd`
cd $workdir


 setenv AMBERHOME /nfs/soft/amber/amber14

rm lig2; mkdir lig2; cd lig2

#cp $workdir/xtal-lig.pdb lig.pdb
#cp $workdir/33443.pdb lig.pdb
cp $workdir/gdp_h.pdb lig.pdb
#sed -i 's/<0> /LIG/g' lig1.mol2

$AMBERHOME/bin/antechamber -i lig.pdb -fi pdb -o lig.ante.mol2 -fo mol2 

$AMBERHOME/bin/antechamber -i lig.ante.mol2 -fi mol2 -o lig.ante.charge.mol2 -fo mol2 -c bcc -at sybyl -nc -3
$AMBERHOME/bin/antechamber -i lig.ante.mol2 -fi mol2  -o lig.ante.pdb  -fo pdb
$AMBERHOME/bin/antechamber -i lig.ante.charge.mol2 -fi mol2  -o lig.ante.charge.prep -fo prepi
$AMBERHOME/bin/parmchk -i lig.ante.charge.prep -f  prepi -o lig.ante.charge.frcmod



Here is the tleap input file:

cat tleap.in | awk '{print " "$0}'
set default PBradii mbondi2
# load the protein force field
source leaprc.ff12SB
# load in GAFF
source leaprc.gaff
# ions
loadamberparams /nfs/soft/amber/amber14/dat/leap/parm/frcmod.ionsjc_tip3p 
loadamberparams /nfs/soft/amber/amber14/dat/leap/parm/frcmod.ionslrcm_hfe_tip3p 

# load ligand and covalent parameters.  
loadamberparams lig/lig.ante.charge.frcmod
loadamberparams lig2/lig.ante.charge.frcmod

loadamberparams thioe.frcmod

loadamberprep lig/lig.ante.charge.mod.prep

loadamberprep lig2/lig.ante.charge.prep

loadamberprep CYB.prep

# load pdb file 
REC = loadpdb rec.pdb
LIG = loadpdb 94F.full_mod_num.pdb 
COF = loadpdb gdp.pdb 
#complex
COM  = combine {REC COF LIG}

# draw bond between CYN and LIG
#bondByDistance COM 2.0
bond COM.172.C18 COM.12.SG 
#bond COM.330.C COM.331.N 
#bond COM.331.C COM.332.N 
#deleteBond COM.331.SG COM.331.C 

#desc COM.331
#desc COM.331.SG
#desc COM.331.C
#desc COM.331.N
#desc COM.501
desc COM.172.C18
desc COM.12.SG 

saveamberparm COM com.leap.prm7 com.leap.rst7

quit

Here is the command to run leap:

$AMBERHOME/bin/tleap -s -f tleap.in > ! tleap.out

Here is the command to submit the miminization to the queue:

cat run.004.pmemd_cuda_min.csh | awk '{print " "$0}'
#setenv AMBERHOME /nfs/soft/amber/amber14

setenv AMBERHOME /nfs/soft/amber/amber14
setenv LD_LIBRARY_PATH ""
#setenv LD_LIBRARY_PATH "/usr/local/cuda-6.0/lib64/:$LD_LIBRARY_PATH"
setenv LD_LIBRARY_PATH "/nfs/soft/cuda-6.5/lib64/:\$LD_LIBRARY_PATH"

cat << EOF1 > ! 01mi.in
01mi.in: minimization with GB
&cntrl
 imin = 1, maxcyc = 10000, ncyc = 500,  ntmin = 1,
 igb=1,
 ntx = 1, ntc = 1, ntf = 1,
 ntb = 0, ntp = 0,
 ntwx = 1000, ntwe = 0, ntpr = 1000,
 cut = 999.9,
 ntr = 1,
 restraintmask = '!@H=', 
 restraint_wt = 0.1,
/
EOF1


#$AMBERHOME/bin/pmemd.cuda -O -i 01mi.in -o 01mi.out -p com.leap.prm7 -c com.leap.rst7 -ref com.leap.rst7 -x 01mi.mdcrd -inf 01mi.info -r 01mi.rst7
#$AMBERHOME/bin/sander -O -i 01mi.in -o 01mi.out -p com.leap.prm7 -c com.leap.rst7 -ref com.leap.rst7 -x 01mi.mdcrd -inf 01mi.info -r 01mi.rst7

set pwd = `pwd`
#cd $pwd
 
cat << EOF > ! qsub.sander.csh
#\$ -S /bin/csh
#\$ -cwd
#\$ -q gpu.q
#\$ -o stdout
#\$ -e stderr

  cd $pwd
  
  $AMBERHOME/bin/pmemd.cuda -O -i 01mi.in -o 01mi.out -p com.leap.prm7 -c com.leap.rst7 -ref com.leap.rst7 -x 01mi.mdcrd -inf 01mi.info -r 01mi.rst7

EOF

  qsub qsub.sander.csh

Visualize the minimized coordinates with Chimera or VMD, you can first convert the files to pdb format with the following commands:

$AMBERHOME/bin/ambpdb -p com.leap.prm7 < 01mi.rst7 > 01mi.pdb
$AMBERHOME/bin/ambpdb -p com.leap.prm7 < com.leap.rst7 > com.leap.pdb