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Hi Fredrik,
Should not mess things up.
As long as the sum of the occupancies of ALIG (LIG would be just fine
since you do not have BLIG) and BPHE does not exceed 1.0 Refmac should
be happy.
You could have a problem if ALIG is too close to APHE though. Checked that?
What if you set all occupancies of BPHE to 0.0?
R.
Quoting Ekström Fredrik <[EMAIL PROTECTED]>:
Dear All,
Thanks for all replies and this interesting discussion!
I'm not really sure that I follow all your answers; probably my description
of the problem is unclear!
To clarify,
In chain A I have one residue (Phe) in two alternative conformations.
In this area, a ligand is accommodated; let us assume for the discussion
that the occupancy is 0.8.
When the ligand is present, the Phe is in a conformation that can coordinate
the ligand. The occupancy of this conformation is 0.8. In the pdb I label
this conformation as APHE and the ligand as ALIG.
Although the resolution is low, I have some density that implicates an
alternative conformation of Phe (similar to apo). This conformation is not
compatible with a ligand binding. The conformation is labelled BPHE in the
pdb, and of course no ligand can be present with Phe is this conformation.
In the refinement, the incompatibility between the apo conformation of Phe
and the ligand mess things up!
Regards
/Fredrik Ekström
-----Ursprungligt meddelande-----
Från: [EMAIL PROTECTED] [mailto:[EMAIL PROTECTED] För Artem
Evdokimov
Skickat: den 17 mars 2006 15:10
Till: 'nic steussy'; [EMAIL PROTECTED]
Ämne: RE: [ccp4bb]: Refmac ligand occupancy refinement
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Hi,
There is an error estimate for every parameter of refined model. However,
for B and O (or N, whatever you want to call occupancy) the parameters are
closely related - therefore the refinement of occupancy is difficult at low
data/parameter ratios - such as encountered at low resolution :)
Realistically, if you see enough of the minor component of the disorder,
even at such a low resolution, then you can and should model it, but I
personally wouldn't bother too much with trying to establish 'accurate'
occupancy at this resolution. In my experience, a guess is about as accurate
as trying to match B-factors.
You can try SHELX - it can refine independent occupancy parameters. At 2.5 A
I would use one B-factor per residue and restrain a whole lot, to make sure
that the 'anatomy' of the structure is realistic. So in the case in
question, there will be one B-factor parameter per residue, and then for the
few residues that are disordered there will be an additional parameter - the
occupancy (N and 1-N for the two components of each pair, so still only one
extra). There shouldn't be an extra B-factor parameter since SHELX can link
B-factors of the disordered pairs. This way you won't be adding too many
parameters (you still have all the extra coordinates to refine, though!).
Free variables FTW*!
Artem
ex-PGP
* - FTW is gamer slang acronym 'For The Win' - meaning 'great'.
-----Original Message-----
From: [EMAIL PROTECTED] [mailto:[EMAIL PROTECTED] On Behalf Of nic
steussy
Sent: Friday, March 17, 2006 5:43 AM
To: [EMAIL PROTECTED]
Subject: Re: [ccp4bb]: Refmac ligand occupancy refinement
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All,
How reasonable is it to estimate the occupancy of a ligand by assuming
it has the same B factors as the adjacent side chain atoms? I don't
mean to pick on Ekstrom, as I have seen this assumption asserted in a
number of structure papers. I was a bit skeptical, especially at modest
(mid 2.0A) resolution. What do you all think?
Nic out
============================
C. Nicklaus Steussy, M.D., Ph.D.
Purdue University
[EMAIL PROTECTED]
============================
Ekström Fredrik wrote:
I'm struggling with a 2.5 Å structure of a protein-ligand complex.
Current R/Rfree is 19/24.
The occupancy of the ligand is approximately 0.80 (estimated by B-factor
matching of neighbouring residues)
--
Dr. Roberto Steiner
Randall Division of Cell and Molecular Biophysics
New Hunt's House
King's College London
Guy's Campus
London, SE1 1UL
Phone +44 (0)20-7848-8216
Fax +44 (0)20-7848-6435
e-mail [EMAIL PROTECTED]
