Hi Dirk,
I disagree with your final sentence. Even if you don't apply NCS
restraints/constraints during refinement, there is a serious risk of NCS
"contaminating" your Rfree. Consider the limiting case in which the
"NCS" is produced simply by working in an artificially low symmetry
space-group (e.g. P1, when the true symmetry is P2): in this case,
putting one symmetry mate in the Rfree set, and one in the Rwork set
will guarantee that Rfree tracks Rwork. The same effect applies to a
large extent even if the NCS is not crystallographic.
Bottom line: thin shells are not a perfect solution, but if NCS is
present, choosing the free set randomly is *never* a better choice, and
almost always significantly worse. Together with multicopy refinement,
randomly chosen test sets were almost certainly a major contributor to
the spuriously good Rfree values associated with the retracted MsbA and
EmrE structures.
Best wishes,
Dean
Dirk Kostrewa wrote:
> Dear CCP4ers,
>
> I'm not convinced, that thin shells are sufficient: I think, in
> principle, one should omit thick shells (greater than the diameter of
> the G-function of the molecule/assembly that is used to describe
> NCS-interactions in reciprocal space), and use the inner thin layer of
> these thick shells, because only those should be completely independent
> of any working set reflections. But this would be too "expensive" given
> the low number of observed reflections that one usually has ...
> However, if you don't apply NCS restraints/constraints, there is no need
> for any such precautions.
>
> Best regards,
>
> Dirk.
>
> Am 07.02.2008 um 16:35 schrieb Doug Ohlendorf:
>
>> It is important when using NCS that the Rfree reflections be selected is
>> distributed thin resolution shells. That way application of NCS should not
>> mix Rwork and Rfree sets. Normal random selection or Rfree + NCS
>> (especially 4x or higher) will drive Rfree down unfairly.
>>
>> Doug Ohlendorf
>>
>> -----Original Message-----
>> From: CCP4 bulletin board [mailto:[log in to unmask]] On Behalf Of
>> Eleanor Dodson
>> Sent: Tuesday, February 05, 2008 3:38 AM
>> To: [log in to unmask] <mailto:[log in to unmask]>
>> Subject: Re: [ccp4bb] an over refined structure
>>
>> I agree that the difference in Rwork to Rfree is quite acceptable at
>> your resolution. You cannot/ should not use Rfactors as a criteria for
>> structure correctness.
>> As Ian points out - choosing a different Rfree set of reflections can
>> change Rfree a good deal.
>> certain NCS operators can relate reflections exactly making it hard to
>> get a truly independent Free R set, and there are other reasons to make
>> it a blunt edged tool.
>>
>> The map is the best validator - are there blobs still not fitted? (maybe
>> side chains you have placed wrongly..) Are there many positive or
>> negative peaks in the difference map? How well does the NCS match the 2
>> molecules?
>>
>> etc etc.
>> Eleanor
>>
>> George M. Sheldrick wrote:
>>> Dear Sun,
>>>
>>> If we take Ian's formula for the ratio of R(free) to R(work) from his
>>> paper Acta D56 (2000) 442-450 and make some reasonable approximations,
>>> we can reformulate it as:
>>>
>>> R(free)/R(work) = sqrt[(1+Q)/(1-Q)] with Q = 0.025pd^3(1-s)
>>>
>>> where s is the fractional solvent content, d is the resolution, p is
>>> the effective number of parameters refined per atom after allowing for
>>> the restraints applied, d^3 means d cubed and sqrt means square root.
>>>
>>> The difficult number to estimate is p. It would be 4 for an isotropic
>>> refinement without any restraints. I guess that p=1.5 might be an
>>> appropriate value for a typical protein refinement (giving an R-factor
>>> ratio of about 1.4 for s=0.6 and d=2.8). In that case, your R-factor
>>> ratio of 0.277/0.215 = 1.29 is well within the allowed range!
>>>
>>> However it should be added that this formula is almost a
>>> self-fulfilling prophesy. If we relax the geometric restraints we
>>> increase p, which then leads to a larger 'allowed' R-factor ratio!
>>>
>>> Best wishes, George
>>>
>>>
>>> Prof. George M. Sheldrick FRS
>>> Dept. Structural Chemistry,
>>> University of Goettingen,
>>> Tammannstr. 4,
>>> D37077 Goettingen, Germany
>>> Tel. +49-551-39-3021 or -3068
>>> Fax. +49-551-39-2582
>>>
>>>
>>>
>
>
> *******************************************************
> Dirk Kostrewa
> Gene Center, A 5.07
> Ludwig-Maximilians-University
> Feodor-Lynen-Str. 25
> 81377 Munich
> Germany
> Phone: +49-89-2180-76845
> Fax: +49-89-2180-76999
> E-mail: [log in to unmask] <mailto:[log in to unmask]>
> *******************************************************
>
>
--
Dean R. Madden, Ph.D.
Department of Biochemistry
Dartmouth Medical School
7200 Vail Building
Hanover, NH 03755-3844 USA
tel: +1 (603) 650-1164
fax: +1 (603) 650-1128
e-mail: [log in to unmask]
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