| Quick navigation: | Home | Site Map || References | Biography || Copyright | Other copyright | Contact us | | |
|
|
Re: [ccp4bb] an over refined structure |
|
CCP4bb navigationCCP4bb <-- 2008 <-- February 2008 <-- 07 February 2008Subject: Re: an over refined structure From: Dean Madden drm0001 {- at -} DARTMOUTH {- dot -} EDU Date: 2008-02-07 of Rwork. However, the whole point of the Rfree is that it is supposed to be independent of the number of parameters you're refining. Simply throwing multiple copies of the model into the refinement shouldn't have affected Rfree, IF IT WERE TRULY "FREE". It was almost certainly NCS-mediated spillover that allowed the multicopy, parameter-driven reduction in Rwork to pull down the Rfree values as well. The experiment is probably not worth the time it would take to do, but I suspect that if MsbA and EmrE test sets had been chosen in thin shells, then Rfree wouldn't have shown nearly the "improvement" it did. Dean Phil Jeffrey wrote: > While NCS probably played a role in the first crystal form of MsbA (P1, > 8 monomers), this is also the one that showed the greatest improvement > in R-free once the structure was correctly redetermined (7% or 14% > depending on which refinement protocols you compare). > > The other crystal form of MsbA and the crystal forms of EmrE didn't have > particularly high-copy NCS (2 dimers, 4 monomers, dimer, 2 tetramers) > and the R-frees were somewhat comparable in all cases (31-36% for the > redetermined structures). > > The *major* source of the R-free suppression in all these cases with the > inappropriate use of multi-copy refinement at low resolution. > > Phil Jeffrey > Princeton > > > Dean Madden wrote: >> 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:CCP4BB@JISCMAIL.AC.UK] On Behalf Of >>>> Eleanor Dodson >>>> Sent: Tuesday, February 05, 2008 3:38 AM >>>> To: CCP4BB@JISCMAIL.AC.UK >>>> 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: kostrewa@lmb.uni-muenchen.de >>> >>> ******************************************************* >>> >>> >> > -- 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: dean.madden@dartmouth.edu CCP4bb navigationCCP4bb <-- 2008 <-- February 2008 <-- 07 February 2008 |
| ProteinCrystallography.org: Copyright 2006-2008 by Quid United Ltd |