AMBER Archive (2008)

Subject: Re: AMBER: Need help... High energies for complex...

From: Waqas Nasir (nasirwaqas1983_at_yahoo.com)
Date: Mon Sep 08 2008 - 11:06:26 CDT

Hi,

Hope every body is doing fine.

Well, first of all thanks a lot for the worthed help. Secondly, just wanted to share what exactly was the problem, and ask if anybody could please explain.

The file that I was using earlier was;

-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
source leaprc.ff99

prot = loadpdb /proteins/frame.pdb.temp
saveamberparm prot /proteins/frame.prmtop.temp /proteins/frame.inpcrd.temp

source leaprc.Glycam_06
sug = loadpdb /sugars/frame.pdb.temp
bond sug.615.C1 sug.618.O2
bond sug.616.O3 sug.618.C1
bond sug.616.C1 sug.617.O1
saveamberparm sug /sugars/frame.prmtop.temp /sugars/frame.inpcrd.temp

comp = combine{prot sug}
saveamberparm comp /complex/frame.prmtop.temp /complex/frame.inpcrd.temp
quit
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------

And now what I am using is;

-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
source leaprc.ff99
source leaprc.Glycam_06

sug = loadpdb /sugars/frame.pdb.temp
bond sug.615.C1 sug.618.O2
bond sug.616.O3 sug.618.C1
bond sug.616.C1 sug.617.O1
saveamberparm sug /sugars/frame.prmtop.temp /sugars/frame.inpcrd.temp

prot = loadpdb /proteins/frame.pdb.temp
saveamberparm prot /proteins/frame.prmtop.temp /proteins/frame.inpcrd.temp

comp = combine{prot sug}
saveamberparm comp /complex/frame.prmtop.temp /complex/frame.inpcrd.temp
quit
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------

The only difference is the order of protein top and crd files generation. It took me several hours to pinpoint that the prmtop file from xleap for protein was different from that of tleap because there in xleap I first generated top and crd for sugar and not the proteins (accidently) which was not the case in tleap. So, for testing purpose I just changed the order in tleap and it worked. Now the results that I have are as follows;

For complex;

NSTEP = 0 TIME(PS) = 0.000 TEMP(K) = 360.57 PRESS = 0.0
 Etot = -10010.3222 EKtot = 8487.1166 EPtot = -18497.4388
 BOND = 1607.6271 ANGLE = 4036.6686 DIHED = 2710.8721
 1-4 NB = 2245.4694 1-4 EEL = 24886.8144 VDWAALS = -5110.6796
 EELEC = -42393.2087 EGB = -6595.4631 RESTRAINT = 0.0000
 ESURF= 114.4610

For proteins;

NSTEP = 0 TIME(PS) = 0.000 TEMP(K) = 360.54 PRESS = 0.0
 Etot = -10271.0692 EKtot = 8398.2944 EPtot = -18669.3636
 BOND = 1564.9753 ANGLE = 3963.4071 DIHED = 2729.7999
 1-4 NB = 2227.4664 1-4 EEL = 24304.9696 VDWAALS = -5059.4524
 EELEC = -42044.1710 EGB = -6470.2727 RESTRAINT = 0.0000
 ESURF= 113.9142

For sugars;

NSTEP = 0 TIME(PS) = 0.000 TEMP(K) = 335.14 PRESS = 0.0
 Etot = 277.0085 EKtot = 79.9191 EPtot = 197.0894
 BOND = 42.6518 ANGLE = 73.2616 DIHED = -18.9470
 1-4 NB = 18.0030 1-4 EEL = 581.8448 VDWAALS = -21.6925
 EELEC = -350.4760 EGB = -131.0059 RESTRAINT = 0.0000
 ESURF= 3.4497

Can any body please explain why is the order important? I really cant understand the logic...
Secondly, after minimization the DIHED values are a bit distorted, I mean they do not accurately sum up to the complex but are close anyways. But the bond and angle values are accurate.
I would appreciate if somebody could comment.

Thanks alot for the help that you people provided me with,
I really do appreciate that..
Thanks once again!
Regards,
Waqas.

----- Original Message ----
From: Carlos Simmerling <carlos.simmerling_at_gmail.com>
To: amber_at_scripps.edu
Sent: Friday, September 5, 2008 12:07:01 PM
Subject: Re: AMBER: Need help... High energies for complex...

no, you cannot conclude anything about binding. all you can conclude
is that there is a problem somewhere in your protocol. anything you
calculate with these will not be correct. you might want to try
working with a calculation that does not require editing of the
molecule or new parameters and see if you can get it to work on a
simpler test case. once you have experience with that, then you might
be able to get this more advanced system to work.

On Fri, Sep 5, 2008 at 5:15 AM, Waqas Nasir <nasirwaqas1983_at_yahoo.com> wrote:
> Hi,
>
> Thanks once again for such a great help.
>
> This time I have used xleap and done every thing with my hand, rather than
> using a script. This time the result that I have got are better but not
> completely correct. I am sorry for the trouble that I am causing...
>
> Here is the single point md for complex;
>
> NSTEP = 0 TIME(PS) = 0.000 TEMP(K) = 360.57 PRESS =
> 0.0
> Etot = -11596.4113 EKtot = 8487.1166 EPtot =
> -20083.5279
> BOND = 205.6103 ANGLE = 1549.3622 DIHED =
> 1658.6999
> 1-4 NB = 2829.8000 1-4 EEL = 25430.2414 VDWAALS =
> -3397.0994
> EELEC = -41648.8133 EGB = -6824.3588 RESTRAINT =
> 0.0000
> ESURF= 113.0299
>
> For sugars;
>
> NSTEP = 0 TIME(PS) = 0.000 TEMP(K) = 335.14 PRESS =
> 0.0
> Etot = 242.2702 EKtot = 79.9191 EPtot =
> 162.3511
> BOND = 15.1725 ANGLE = 47.3410 DIHED =
> -27.5845
> 1-4 NB = 26.3282 1-4 EEL = 592.4065 VDWAALS =
> -16.3978
> EELEC = -374.1894 EGB = -104.0077 RESTRAINT =
> 0.0000
> ESURF= 3.2822
>
> For proteins;
>
> NSTEP = 0 TIME(PS) = 0.000 TEMP(K) = 360.54 PRESS =
> 0.0
> Etot = -8302.0894 EKtot = 8398.2944 EPtot =
> -16700.3838
> BOND = 190.4378 ANGLE = 1485.0230 DIHED =
> 5266.2811
> 1-4 NB = 2803.4718 1-4 EEL = 24837.8348 VDWAALS =
> -3357.5193
> EELEC = -41233.4669 EGB = -6804.6020 RESTRAINT =
> 0.0000
> ESURF= 112.1559
>
> The energies are still ridiculous... but the bond energy is accurately
> additive where as the angles are almost. But the dihedrals are completely
> out. This time I have done every thing with hand so I know that the prmtop
> and inpcrd files are fine. What option can you think of now... to debug...
> should I go back to the way I created the sugars and everything... I just
> want to confirm a couple of things;
>
> 1. Is the protocol legitimate and proper (The way that I am calculating
> single point energy for each frame)?
> 2. Do you still think that there might be some problem in prmtop and inpcrd
> files (looking at the results)?
> 3. Do you think that these results might reflect the improper state of
> binding of ligand and the proteins, I mean can I conclude from these results
> that the sugar in question DOES NOT bind to proteins at all!!! I am running
> a small 200 ps md without restraints on this complex to see if the ligand
> remains in the binding pocket or flies off...
>
> I would really appreciate if you could comment please... I am new to amber
> and had just started to enjoy it until this happened...
>
> Thanks, and extremely sorry for the trouble that I am causing...
> Regards,
> Waqas.
>
> ----- Original Message ----
> From: Carlos Simmerling <carlos.simmerling_at_gmail.com>
> To: amber_at_scripps.edu
> Sent: Thursday, September 4, 2008 6:21:42 PM
> Subject: Re: AMBER: Need help... High energies for complex...
>
> if the coordinates are identical, and there is no covalent link, then
> the bond/angle/dihedral energies should be additive. in your case they
> are far from that - look at the bonds.. if there are no bonds between
> protein and sugar, how can the bond energies not sum up? the only
> conclusion I can make is that the parameters in the individual prmtop
> files do not match those in the prmtop for the complex, or perhaps you
> are not really using the same protein coordinates for the complex and
> isolated protein, or for complex and isolated sugar. I don't really
> have any way of knowing how that might have happened.
>
> On Thu, Sep 4, 2008 at 12:14 PM, Waqas Nasir <nasirwaqas1983_at_yahoo.com>
> wrote:
>>
>> The last mail that I sent you it had exactly the same coordinates in the
>> complex and the separate subunits. I just pasted the coordinates from main
>> file in two separate files for protein and sugar subunits.
>>
>> I will re check with the prmtop and crd files but I doubt if there is
>> anything that I could do... its just 3 bond commands and one saveamberparm
>> command. I dont have enough room to debug. And yes there is no covalent
>> linkage between the sugar and protein.
>> Do you think that its dead sure that something is different in top and crd
>> files ???
>> Do you think, moreover, that combine command might have done something
>> wrong... just a thought...
>>
>> Thanks,
>> Waqas.
>>
>> ----- Original Message ----
>> From: Carlos Simmerling <carlos.simmerling_at_gmail.com>
>> To: amber_at_scripps.edu
>> Sent: Thursday, September 4, 2008 6:41:38 PM
>> Subject: Re: AMBER: Need help... High energies for complex...
>>
>> are the coordinates you are using for the complex exactly the same as
>> for the protein and sugar alone (the energies you posted earlier)?
>> if not, then you really can't use this to debug your energies. they
>> must be identical (but separate files).
>> if yes, then there is something wrong in the prmtop files and you will
>> need to go through your procedure for generating the separate files
>> and make sure they are the same.
>> all of this assumes no covalent link between sugar and protein.
>>
>> On Thu, Sep 4, 2008 at 11:22 AM, Waqas Nasir <nasirwaqas1983_at_yahoo.com>
>> wrote:
>>> Hi,
>>>
>>> Well, thanks a lot for the worthed help.
>>>
>>> Let me explain the whole procedure that I have taken up, I have some
>>> doubts
>>> in the way I have separated the files and the first thing that came into
>>> my
>>> mind after these results was exactly that the top files are not
>>> corresponding to what is in the results.
>>>
>>> Anyways, the sugar that I have is a tetra saccharide. I first constructed
>>> the sugar unit in xleap and added 3 bonds which were missing when I
>>> imported the pdb file into the xleap with glycam_06 force field (The
>>> reason
>>> being not able to find the exact residues in the glycam06 force field as
>>> are
>>> in the pdb file). Along side that I had a protein unit from the part of
>>> the
>>> pdb file which contained protein. I combined both units in xleap by
>>> combine
>>> command and then generated the prmtop and inpcrd files for the main
>>> complex
>>> on which I had all the minimization and md runs going on.
>>>
>>> Secondly the script for generating frames generated 250 pdb files
>>> corresponding to the 250 frames that I had in the trajectory. I again
>>> repeated the same procedure using tleap and constructed a sugar subunit
>>> and
>>> a protein subunit for each frame and generated top and crd files
>>> separately
>>> for the two subunits in each frame. These files were then used for
>>> subsequent single point energy calculations.
>>>
>>> Do you see mistakes in principle of the approach that has been taken up
>>> here???
>>>
>>> Awaiting your response,
>>> Thanks a lot,
>>> Waqas.
>>>
>>> ----- Original Message ----
>>> From: David A. Case <case_at_biomaps.rutgers.edu>
>>> To: amber_at_scripps.edu
>>> Sent: Thursday, September 4, 2008 5:36:37 PM
>>> Subject: Re: AMBER: Need help... High energies for complex...
>>>
>>> On Thu, Sep 04, 2008, Waqas Nasir wrote:
>>>>
>>>> Well, I have tried straight single point md on the complex,sugar and
>>>> protein
>>>> separately, and what I have found is as follows;
>>>
>>> This is not what you should do, at least until you understand what is
>>> going on. My suggestion (and Carlos'):
>>>
>>> Make a single prmtop file for the complex. Use this prmtop file for
>>> a single point calculation with two sets of coordinates: one with the
>>> ligand
>>> (sugar) in its binding location, and a second set of coordinates where
>>> you
>>> manually move the ligand to someplace distant from the protein, (but keep
>>> the
>>> protein coordinates identical to what they were in the complex.
>>>
>>> Once you see and understand how that works, you will be ready to try and
>>> interpret other experiments.
>>>
>>> ...regards...dac
>>>
>>> -----------------------------------------------------------------------
>>> The AMBER Mail Reflector
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>>>
>>>
>>
>>
>>
>> --
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