If it is crystal packing disorder (F's added instead of I's), the switches
between the alternative conformations need to be very frequent, to be within
coherent range, so I would asume that the alternative conformations will be
present in equal proportions. Still the alternatives need to be modeled somehow
and if this can be conveniently done in a lower symmetry spacegroup this would
not artificially lower the free R-factors. As Phoebe mentioned, ignoring the
higher symmetry relations and repicking the free Rflags at lower symmetry would
lead free reflections to be linked to the working set, leading to too low Rfree
values. However, with perfect packing disorder, no extra information would be
gained by reprocessing in lower symmetry (in contrast to cases with pseudo
symmetry).
My 2 cents,
Herman
-----Original Message-----
From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Phoebe A.
Rice
Sent: Tuesday, March 19, 2013 4:49 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Strange density in solvent channel and high Rfree
oops, I should have expanded my comments to include the sort of funky lattice
order-disorder Zbyszek so cleverly diagnosed. Scratch that "perfect twinning"
comment in my last message.
________________________________________
From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Phoebe A. Rice
[pr...@uchicago.edu]
Sent: Tuesday, March 19, 2013 10:34 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Strange density in solvent channel and high Rfree
Hi Zbyszek,
If the issue is perfect twinning, I agree - good point!
But you don't want to confuse people who simply have nearly-but-not-quite
crystallographic symmetry (OK, I'm being a bit pedagogical here, but a lot of
newbies read the BB). We had a case of P31 that was so close to P61 we
actually solved the molecular replacement problem in P61, then expanded it back
and re-rigid-bodied it. We've played similar games with translational
pseudo-symmetry (ignoring the weak spots at first). In cases like that it is
important to properly reprocess the data in the lower symmetry space group (or
smaller unit cell) because there is real information in those small
differences. However, the point about Rfree holds for twinning or rotational
pseudo-symmetry: the Rfree flags should be expanded by the xtal symmetry
operators, not re-picked in the lower symmetry space group.
Phoebe
++++++++++++++++++++++++++++++++++++++++++
Phoebe A. Rice
Dept. of Biochemistry& Molecular Biology The University of Chicago
773 834 1723; pr...@uchicago.edu
http://bmb.bsd.uchicago.edu/Faculty_and_Research/
http://www.rsc.org/shop/books/2008/9780854042722.asp
________________________________________
From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Zbyszek
Otwinowski [zbys...@work.swmed.edu]
Sent: Tuesday, March 19, 2013 9:37 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Strange density in solvent channel and high Rfree
It is a clear-cut case of crystal packing disorder. The tell-tale sign is that
data can be merged in the higher-symmetry lattice, while the number of
molecules in the asymmetric unit (3 in P21) is not divisible by the higher
symmetry factor (2, by going from P21 to P21212).
From my experience, this is more likely a case of order-disorder than
merohedral twinning. The difference between these two is that structure factors
are added for the alternative conformations in the case of order-disorder,
while intensities (structure factors squared) are added in the case of
merohedral twinning.
Now an important comment on how to proceed in the cases where data can be
merged in a higher symmetry, but the structure needs to be solved in a lower
symmetry due to a disorder.
!Such data needs to be merged in the higher symmetry,assigned R-free flag, and
THEN expanded to the lower symmetry. Reprocessing the data in a lower symmetry
is an absolutely wrong procedure and it will artificially reduce R-free, as the
new R-free flags will not follow data symmetry!
Moreover, while this one is likely to be a case of order-disorder, and these
are infrequent, reprocessing the data in a lower symmetry seems to be
frequently abused, essentially in order to reduce R-free. Generally, when data
CAN be merged in a higher symmetry, the only proper procedure in going to a
lower-symmetry structure is by expanding these higher-symmetry data to a lower
symmetry, and not by rescaling and merging the data in a lower symmetry.
Zbyszek Otwinowski
Dear all,
We have solved the problem. Data processing in P1 looks better (six
molecules in ASU), and Zanuda shows a P 1 21 1 symmetry (three
molecules in ASU), Rfactor/Rfree drops to 0.20978/0.25719 in the first
round of refinement (without put waters, ligands, etc.).
Indeed, there were one more molecule in ASU, but the over-merged data
in an orthorhombic lattice hid the correct solution.
Thank you very much for all your suggestions, they were very important
to solve this problem.
Cheers,
Andrey
2013/3/15 Andrey Nascimento<andreynascime...@gmail.com>
*Dear all,*
*I have collected a good quality dataset of a protein with 64% of
solvent in P 2 21 21 space group at 1.7A resolution with good
statistical parameters (values for last shell: Rmerge=0.202;
I/Isig.=4.4; Complet.=93% Redun.=2.4, the overall values are better
than last shell). The structure solution with molecular replacement
goes well, the map quality at the protein chain is very good, but in
the final of refinement, after addition of a lot of waters and other
solvent molecules, TLS refinement, etc. ...
the Rfree is a quite high yet, considering this resolution
(1.77A).(Rfree=
0.29966 and Rfactor= 0.25534). Moreover, I reprocess the data in a
lower symmetry space group (P21), but I got the same problem, and I
tried all possible space groups for P222, but with other screw axis I
can not even solve the structure.*
*A strange thing in the structure are the large solvent channels with
a lot of electron density positive peaks!? I usually did not see too
many peaks in the solvent channel like this. This peaks are the only
reason for these high R's in refinement that I can find. But, why are
there too many peaks in the solvent channel???*
*I put a .pdf file (ccp4bb_maps.pdf) with some more information and
map figures in this link:
https://dl.dropbox.com/u/16221126/ccp4bb_maps.pdf*
*
*
*Do someone have an explanation or solution for this?*
* *
*Cheers,*
*Andrey*
Zbyszek Otwinowski
UT Southwestern Medical Center at Dallas
5323 Harry Hines Blvd.
Dallas, TX 75390-8816
Tel. 214-645-6385
Fax. 214-645-6353
