Dear all,

I'm trying to keep some part (ligand) fixed in space using the restraint features 9, 10 and 11 and the restraint type 34. Unfortunately, I was not quite successful with this approach (outlined below). Can anybody give me an advice, how to proceed correctly (correct syntax for the restraints, other approach giving the same result, ...)? Any help is highly appreciated!

Thanks in advance,

Frank

My approach was: # generating a restraint file from the .ini file of a corresponding run, which included the ligand (atom-ids: 1936-1958) using an awk script:

.ini-file:

HETATM 1936 C5 LAE 257 46.483 11.136 15.931 1.00 -0.04 4SG1940 HETATM 1937 C4 LAE 257 45.927 12.329 16.596 1.00 0.07 4SG1941 [...] HETATM 1958 C16 LAE 257 42.378 10.076 17.821 1.00 -0.02 4SG1962 END

.rsr-file:

MODELLER5 VERSION: MODELLER FORMAT R 3 1 9 34 1 2 0 1936 46.483 0.0 R 3 1 10 34 1 2 0 1936 11.136 0.0 R 3 1 11 34 1 2 0 1936 15.931 0.0 R 3 1 9 34 1 2 0 1937 45.927 0.0 R 3 1 10 34 1 2 0 1937 12.329 0.0 R 3 1 11 34 1 2 0 1937 16.596 0.0 [...] R 3 1 9 34 1 2 0 1958 42.378 0.0 R 3 1 10 34 1 2 0 1958 10.076 0.0 R 3 1 11 34 1 2 0 1958 17.821 0.0

My idea was to constrain the x, y or z coordinate (feature 9, 10, 11) of the ligand with a harmonic potential (3) written into restraint type 34 and applied on 1 atom (using 2 parameters: the coordinate and the standard deviation). I've used both 0.0 and higher values as standard deviations to fix the atom position absolutely or to allow some movement, however both was unsuccessful. Perhaps the whole syntax is false???

However, modeller7v7 does not complain about the restraints (as can be seen from the .log file), which I read in as part of my rewritten 'special_restraints' section:

.top-file:

subroutine routine = 'special_restraints' [...] read_restraints file = '/home/literatur/boeckler/D4proj/top/LIG_posconst_new2.rsr', add_restraints = on [...] return end_subroutine

.log-file:

[...] make_re_422_> Number of previous, current restraints : 20593 20593 make_re_423_> Number of previous, current selected restraints: 20327 20327 [...] TOP_________> 218 750 CALL ROUTINE = 'special_restraints' [...] TOP_________> 223 761 READ_RESTRAINTS FILE = '/home/literatur/boeckler/D4proj/t; op/LIG_posconst_new2.rsr', ADD_RESTRAINTS = ON

openf5__224_> Open 11 OLD SEQUENTIAL /home/literatur/boeckler/D4proj/top/LIG_posconst_new2.rsr openf5__224_> Open 11 OLD SEQUENTIAL /home/literatur/boeckler/D4proj/top/LIG_posconst_new2.rsr rdcsr2__307_> Number of restraints read : 69 Number of excluded pairs read: 0 Number of pseudo atoms read : 0

Dynamically allocated memory at amprmcns [B,kB,MB]: 18611512 18175.305 17.749 openf5__224_> Open 11 OLD SEQUENTIAL /home/literatur/boeckler/D4proj/top/LIG_posconst_new2.rsr openf5__224_> Open 11 OLD SEQUENTIAL /home/literatur/boeckler/D4proj/top/LIG_posconst_new2.rsr rdcsr2__307_> Number of restraints read : 69 Number of excluded pairs read: 0 Number of pseudo atoms read : 0 rdcsrs__304_> Restraints in memory, selected restraints: 20662 20396 Explicitly excluded atom pairs in memory : 0 Pseudo atoms in memory : 0 [...]

Thus, it seems that the restraints have been read in successfully and no errors were reported. However, having a look at the output file shows that the ligand has been "exploding", spreading its atoms all in space.

.pdb-file:

HETATM 1936 C5 LAE 257 127.363 17.224 57.224 1.00 57.83 4SG1940 HETATM 1937 C4 LAE 257 50.824 -79.520 -76.644 1.00 57.83 4SG1941 [...] HETATM 1958 C16 LAE 257 66.463 23.642 160.673 1.00 57.83 4SG1962 END