ram bio wrote:
Dear Justin,

Thanks for the options suggested.

I have used -princ and rotate 0 0 90 options with editconf and was
able to place the protein vertically and at the centre of the DPPC
bilayer (128 + 3655) from the site provided in the tutorial, but still
the part of protein is outside the DPPC layer (examined system.gro
(protein newbox +dppc128.gr0) in VMD). The commands used were


This isn't necessarily a problem. So your protein sticks out - that will probably be the case in most membrane protein systems.

editconf -f protein_processed.gro -o protein_newbox.gro  -c -box
6.41840 6.44350 6.59650 -rotate 0 0 90


Depending on the degree to which the protein is protruding out of the bilayer, you may have to adjust the z dimension of the box, as I've suggested before.

cat protein_newbox.gro dppc128.gro > system.gro


so now, i tried to increase the size of the box by redoing it and
using the command

editconf -f protein_processed.gro -o protein_newbox.gro -c -box
6.41840 6.44350 6.59650 -d 1.0 -rotate 0 0 90 -bt cubic


I don't know what effect this will have. It is contradictory to specify a non-cubic box by assigning the vectors explicitly, then using -bt cubic. I doubt you're getting what you think you are, but I don't know which option has precendence, -box or -bt. Either way, you're not doing anything to increase the box size. Also note that the box size in dppc128.gro would have to be similarly manipulated to actually get a larger box.

cat protein_newbox.gro dppc128.gro > system1.gro

I also used various box types like dodecahedron , octahedron but was

These boxes make no sense for a square cross-section of a membrane.

unable increase the size of the box and fix the protein in the centre
of the dppcbilayer (128 + 3655 H2O). Please suggest me how to correct
my command so that all the protein lies in the dppcbilayer and is in
the  centre and can proceed to inflategro steps, also do i need to use
a dppclayer with more lipids like 256 lipid bilayer.

Well, using editconf properly will help. Define a proper box with -box, don't just keep copying the one I use in the tutorial. The x and y components may stay the same, but you may have to increase the z vector.

The same process applies with any bilayer, even if it has 256 lipids.

-Justin


Thanks,

Ram

On Tue, Oct 6, 2009 at 5:39 PM, Justin A. Lemkul <jalem...@vt.edu> wrote:

ram bio wrote:
Dear Justin,

As suggested, when i reexamined the system.gro (protein_newbox +
dppc128) one of the ends of the problem (as i think) i.e. few
aminoacids of protein were beyond the water surface of the bilayer,
probably this may be the reason for the presence of water molecules to
the side of the bilayer when solvated and also lack of minimization
during inflategro step. One more thing, I am inserting the protein in
Indeed, if your system doesn't actually fit within the unit cell you've
defined, you're in for trouble.  Always look at your output!

lipid bilayer by orienting it using your KALP peptide (.pdb) as
reference in VMD and later using editconf to centre the protein using
option i.e. -c -box 6.41840 6.44350 6.59650, otherwise if i just use
editconf command, it is orienting horizontally instead of vertically,
I don't fully understand what you're doing.  You also may not be able to use
the exact same box that I defined in the tutorial (the original DPPC box).
 If you've got pieces of your protein protruding out of the unit cell, then
you'll need to define a suitably-sized box.  Horizontal vs. vertical issues
can be solved by editconf -rotate.

-Justin

i can do the orientation manually using vmd but it is difficult and
iam unable to orient it exactly in the centre and vertical in the dppc
bilayer.

Please suggest some corrections as I am going to reorient and position
it in the bilayer and redo  the inflategro procedure.

Thanks

Ram

On Tue, Oct 6, 2009 at 4:42 PM, Justin A. Lemkul <jalem...@vt.edu> wrote:
ram bio wrote:
Dear Justin,

Thanks for the advice. I am using the DPPC 128 lipid bilayer from D.
Peter Tieleman website, and the nvt.mdp file and the nvt.log files are
as follows as in your tutorial:
<snip>

Constraining the starting coordinates (step 0)

Constraining the coordinates at t0-dt (step 0)
RMS relative constraint deviation after constraining: 9.42e-04
Initial temperature: 503.557 K

Started mdrun on node 0 Thu Jun 25 11:30:30 2009

         Step           Time         Lambda
            0        0.00000        0.00000

Grid: 9 x 9 x 9 cells
Large VCM(group Protein_DPPC):    -50.08205,     97.99061,
16.32530, Temp-cm:  2.50669e+05
Long Range LJ corr.: <C6> 2.0307e-03
Long Range LJ corr.: Epot   -1862.02, Pres:    -184.12, Vir:    1862.02
 Energies (kJ/mol)
        Angle       G96Angle    Proper Dih. Ryckaert-Bell.  Improper
Dih.
  1.48814e+04    8.19090e+03    8.43857e+03    6.38969e+03
 2.93352e+03
        LJ-14     Coulomb-14        LJ (SR)  Disper. corr.   Coulomb
(SR)
  4.09831e+03    5.49186e+04    7.87055e+09   -1.86202e+03
-1.48414e+05
 Coul. recip. Position Rest.      Potential    Kinetic En.   Total
Energy
 -1.53985e+05    9.50084e+00    7.87034e+09    3.08099e+17
 3.08099e+17
 Conserved En.    Temperature Pressure (bar)  Cons. rmsd ()
  3.08099e+17    2.31982e+15    1.01551e+16    7.25066e+04

The information shown here indicates very strongly that you have severe
atomic overlap in your starting structure.  This is probably from the
InflateGRO minimization that did not converge appropriately.  Your
potential
energy is astronomically high, as well as factors like temperature, and
thus
kinetic energy.  The latter are related to trying to constrain an
inappropriate starting structure.

I would suggest going back to the initial construction stage, figuring
out
why that minimization didn't converge, and start over from there.
 Plowing
ahead when you get unfavorable results is a recipe for LINCS warnings.

-Justin

Please diagnose the information and suggest.

Thanks

ram


On Tue, Oct 6, 2009 at 4:12 PM, Justin A. Lemkul <jalem...@vt.edu>
wrote:
ram bio wrote:
Dear Gromacs Users,

I have inserted the protein in lipid bilayer and performed Inflategro
I am able to reach the required area per lipid after certain
iterations but was unable to get the standard Epot and Fmax values
that is negative and to the power of 5 or 6 and Fmax less than 1000
You won't get that magnitude of Epot without water.  If you can't reach
Fmax
< 1000, you shouldn't just plow ahead.  Analyze where the problem is,
because it is unlikely to go away by magic!

during the last minimization , later i solvated the protein using a
vanderwaal radii for carbon as 0.375, i found some water molecules not
in the core of the lipid layer but to the sides, as they were not in
the centre i ionated the complex with 13 chloride ions as the charge
Did you get rid of these waters?  You can always try tweaking the entry
in
vdwradii.dat for C.  As far as those on the sides are concerned, it
sounds
like the box you've created is too large for the lipids.  I wouldn't
use
this system, because you'll waste a huge amount of time hoping it
equilibrates right.

shown was non-zero total intergral charge 1.30000e+01, then i minized
the ionized complex with the minim.mdp file in as per justin tutorial,
i am following all the mdp files as per the tutorial till now, and was
able to obtain Potential Energy  = -1.8947278e+05
Maximum force     =  9.1642163e+02 on atom 7647
Norm of force     =  5.0845932e+01

That looks fine, but I still think there is an underlying problem in
your
InflateGRO construction step.

and then i created the index file ane while running the nvt
equilllibrqtion i am getting

Step 0, time 0 (ps)  LINCS WARNING
relative constraint deviation after LINCS:
rms 0.007117, max 0.443345 (between atoms 6311 and 6312)
bonds that rotated more than 30 degrees:
 atom 1 atom 2  angle  previous, current, constraint length

The classic case of "blowing up."  See either my Advanced
Troubleshooting
page in the tutorial, or
http://www.gromacs.org/Documentation/Terminology/Blowing_Up.  If you
want
more specific advice, you'll have to provide information at least about
what
type of lipid you're using, and what you have in your .mdp file.

Can any body suggest me how to rectify the defect and is it the
problem with compliation as i am using gromacs 4.0.3 or the memory
space or my running the job.
Well, first off I'd recommend always using the most current version of
the
software, not that it's likely to impact your system, but just in
general.
 This is a problem reported to this list almost daily, so please also
check
the archives.  There are literally thousands of posts regarding LINCS
warnings and unstable systems.

-Justin

Thanks

Ram
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--
========================================

Justin A. Lemkul
Ph.D. Candidate
ICTAS Doctoral Scholar
Department of Biochemistry
Virginia Tech
Blacksburg, VA
jalemkul[at]vt.edu | (540) 231-9080
http://www.bevanlab.biochem.vt.edu/Pages/Personal/justin

========================================
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--
========================================

Justin A. Lemkul
Ph.D. Candidate
ICTAS Doctoral Scholar
Department of Biochemistry
Virginia Tech
Blacksburg, VA
jalemkul[at]vt.edu | (540) 231-9080
http://www.bevanlab.biochem.vt.edu/Pages/Personal/justin

========================================
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interface
or send it to gmx-users-requ...@gromacs.org.
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--
========================================

Justin A. Lemkul
Ph.D. Candidate
ICTAS Doctoral Scholar
Department of Biochemistry
Virginia Tech
Blacksburg, VA
jalemkul[at]vt.edu | (540) 231-9080
http://www.bevanlab.biochem.vt.edu/Pages/Personal/justin

========================================
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--
========================================

Justin A. Lemkul
Ph.D. Candidate
ICTAS Doctoral Scholar
Department of Biochemistry
Virginia Tech
Blacksburg, VA
jalemkul[at]vt.edu | (540) 231-9080
http://www.bevanlab.biochem.vt.edu/Pages/Personal/justin

========================================
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