Dear Amber users,
First of all thank you very much for the replies from Dr. David Case and Dr.
Samantha Hughes However, I still have some further questions. After creating the
.pdb file from the .rst file in Carnal ( I understand that Ambpdb can also do
this), I then use Vi improve (a text editor) to change GLU264 to GLN264, save the
modified .pdb file and then load it up using loadpdb in Xleap. The .pdb file
created by carnal contains all the TER cards between each water molecule. When
Xleap load this .pdb file it gives the same error message as before:
>
> > mutant2 = loadpdb "./mutantGLN_b4_xleap2.pdb"
> Loading PDB file: ./mutantGLN_b4_xleap2.pdb
> -- residue 10240: duplicate H1 atoms (total 10)
> -- residue 10240: duplicate H2 atoms (total 10)
> -- residue 10240: duplicate O atoms (total 10)
> -- residue 10496: duplicate H1 atoms (total 10)
> -- residue 10496: duplicate H2 atoms (total 10)
> -- residue 10496: duplicate O atoms (total 10)
> -- residue 10752: duplicate H1 atoms (total 10)
> -- residue 10752: duplicate H2 atoms (total 10)
> -- residue 10752: duplicate O atoms (total 10)
> -- residue 11008: duplicate H1 atoms (total 10)
> -- residue 11008: duplicate H2 atoms (total 10)
> -- residue 11008: duplicate O atoms (total 10)
> -- residue 11264: duplicate H1 atoms (total 10)
> -- residue 11264: duplicate H2 atoms (total 10)
>
> and so on...
>
> ATOM NAMES IN EACH RESIDUE MUST BE UNIQUE:
> (same-name atoms are reduced to a single atom)
>
> total atoms in file: 96182
When I save this new pdb file created by Xleap using savepdb, the pdb file obtained
only contained 58834 atoms as compared to 96182 atoms from the previous native pdb
file. Some water molecules were deleted in the new mutant.pdb created by Xleap and
MG ions and Na + ions are now in the end of the pdb file instead of the water
molecules as in the native pdb file. When I view the new mutant.pdb file using edit
in xleap, the cubic solvent box now disappears and water molecules are scattering
around the protein solute.
I did try just changing GLU264 to GLN 264 only by changing OE2 to NE2 without
adding 1HE2 and 2HE2 ( I was hoping that Xleap will automatically add these H
atoms), the result pdb file is the same as adding all the GLN residues in place of
GLU in Vi editor i.e. Xleap still gives the same error as above and deletes some
water molecules.
The end of the .rst file of the native protien (using tail -n50 native.rst ) is as
below;
0.3001629 -1.1166741 -0.6605267 -0.2038014 -0.2880406 -0.1830363
-0.0657320 -0.3995468 -0.0811690 -0.1701587 -0.1748538 -0.0692379
-0.1212022 0.0662336 -0.1707823 -0.5347934 0.3395682 -0.3010556
-0.5119471 0.7187285 -0.0372361 -0.0661226 0.0552852 0.1672029
0.2864035 -0.0419988 -0.6367714 -0.5809506 -0.4151232 0.5673318
0.1715268 -0.1473862 0.0623606 -0.0868227 -0.4496734 1.7255690
-1.1333923 0.7800585 -0.4162019 0.1118782 -0.2190952 -0.3884225
-0.1654280 -0.6467546 -0.4936489 -0.3272698 -0.9377690 -0.7325281
0.0717179 -0.1423362 0.2333974 0.5964171 -0.2122841 0.3908791
0.4703812 -0.4800672 -0.0933463 0.1605087 0.0818827 0.2352233
-0.3884374 -0.1792472 -0.2437926 -1.1743770 0.6237560 -0.4007575
0.0285637 0.2736953 0.0946145 -0.2237768 -0.0083521 0.0672300
0.2309629 -0.2537442 0.7138360 0.2009422 -0.0171013 -0.1140876
-0.3605485 0.0610312 0.0913918 0.2419084 0.2445966 -0.1801767
0.0674050 -0.1502339 -0.2054874 0.8682075 0.3474161 -0.5254936
-0.9547344 -0.6934571 0.3191621 0.1020523 -0.4387149 0.1259357
0.6532144 0.4609473 -0.7988763 -0.7945644 1.0331419 -0.2517538
0.1707389 -0.1054298 0.0526729 -0.8066962 0.8557983 0.2840782
0.9702327 -0.0163874 -0.6870759 0.1152681 -0.0152207 -0.0073999
-0.4651218 -0.1447466 0.0928135 0.9184508 -0.1398275 -0.4085805
108.9398670 101.1957914 86.2799423 90.0000000 90.0000000 90.0000000
>
The box info is in the last line of the restart file. I'm not quite sure what I
need to do to use the setBox command in Xleap as Dr. David Case suggested when I
cannot get the right number of water molecules in my mutant.pdb file to begin with.
It said in the AMBER6 Xleap manual that setBox will not add any solvent to the
system. If anyone has come across this problem before, could you please give me any
suggestion or advice? I don't really know what I did wrong here.
Thank you very much for your concern.
Sally
"David A. Case" wrote:
> On Fri, Jul 19, 2002, Salinthip Thipayang wrote:
>
> > I am trying to change an amino acid residue in the active site of a protein
> > The next step is to modify GLU in the active site to GLN and then compare the
> > binding free energies.
> >
> > I am thinking of the following approaches.
> >
> > 1) Create the PDB file from the restart file obtained from the MD run using
> > CARNAL
> > 2) in XLEAP, loadpdb and then change the GLU264 to GLN 264
>
> Much easier, I think: use a text editor to modify the pdb file to change GLU
> to GLN,, change the atoms be what GLN would expect. Then use loadpdb to
> read that into LEaP. And, use ambpdb to get the pdb file; this will ensure
> that TER cards are placed between all of the waters. (That may have been
> the origin of the "duplicate atoms" problem...it's hard to tell without more
> information).
>
> You will need to use a setBox command in leap to make sure the box info
> is in the new prmtop file; get the box info from your last output, or from
> the bottom of the restart file.
>
> > 3) run belly MD first only allow the GLN264 to move and fix other residues and
> > water and then run another belly MD allowing all the residues to move except
> > the ligand so that the ligand can find the optimal structure with all the
> > residues.
> > 4) then run full MD for the whole system.
>
> This sounds OK.
>
> > >
> > > > mutant2 = loadpdb "./mutantGLN_b4_xleap2.pdb"
> > > Loading PDB file: ./mutantGLN_b4_xleap2.pdb
> > > -- residue 10240: duplicate H1 atoms (total 10)
> > > -- residue 10240: duplicate H2 atoms (total 10)
> > > -- residue 10240: duplicate O atoms (total 10)
>
> You should(?) be able to figure out what is going on by looking at the pdb
> file. For some reason, LEaP thinks there are 10 H1 atoms in a single residue.
> Make sure that the residue numbers are unique. Since you have more than
> 10000 residues, this may be a problem with residue numbering: check that
> you don't have "*****" or somehting like that as a residue number in the
> pdb file.
>
> (Aside: I don't think your problem has anything to do with the fact that
> GLN has more atoms than GLU).
>
> (Another aside: depending on how detailed an answer you want, you might
> consider the following: just "neutralize" the GLU by changing the charges
> so they add up to zero for the residue, rather than to -1.) Then you don't
> need to change anything except the few charges, which you could even do in
> by hand in the prmtop file. This would capture to a large extent the
> electrostatic change of this mutation; of course it would miss any details of
> hydrogen bond donor properties of the NH2 group in GLN.)
>
> ..hope this helps....good luck...dac
>
> --
>
> ==================================================================
> David A. Case | e-mail: case_at_scripps.edu
> Dept. of Molecular Biology, TPC15 | fax: +1-858-784-8896
> The Scripps Research Institute | phone: +1-858-784-9768
> 10550 N. Torrey Pines Rd. | home page:
> La Jolla CA 92037 USA | http://www.scripps.edu/case
> ==================================================================
--
-------------------------------------------------------------------------------
Salinthip Thipayang (Miss)
PhD Research Student
Biological and Biophysical Chemistry Research Group
Chemistry Department
Imperial College of Science, Technology and Medicine
Exhibition Road
London SW7 2AY
The United Kingdom
Tel: +44(0)207 5945851
Fax: +44(0)207 5945845
Email: salinthip.thipayang_at_ic.ac.uk
--------------------------------------------------------------------------------
Received on Tue Jul 23 2002 - 07:46:20 PDT
