Re: [ccp4bb] Question about R/Rfree value difference

- Protein crystallography

Main steps:

- Protein purification
- Crystallisation

Special:

- Programs for crystallography
- X-ray detectors

Basic tutorials:

   - Chemistry
   - Protein
   - Peptide
   - Amino Acids

Xtal community:

- CCP4BB

CCP4bb navigation

CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

Previous message:
Subject: Re: Question about R/Rfree value difference
From: Tim Gruene tg {- at -} SHELX {- dot -} UNI-AC {- dot -} GWDG {- dot -} DE
Date: 2010-07-08

Next message:
Subject: Re: Question about R/Rfree value difference
From: Tim Gruene tg {- at -} SHELX {- dot -} UNI-AC {- dot -} GWDG {- dot -} DE
Date: 2010-07-08

Subject: Re: Question about R/Rfree value difference
From: Anastassis Perrakis a {- dot -} perrakis {- at -} NKI {- dot -} NL
Date: 2010-07-08

I do agree with Tim's reasoning in general, but as Pavel also implied
by offering the statistics,
I would not be worried about the difference, but by the unreasonably
high absolute value of Free R for 2.0 A resolution.

I do not think that its simple 'over-fitting' and my worry would not
be just the very high difference between
R/Rfree, but simply the high Rfree. I would more or less bet that if
you check your model in
the MolProbity server (http://molprobity.biochem.duke.edu/) it will
also have appalling geometry scores(*)

Here is what I would suspect and what I would do to check what is wrong:

-very incomplete or badly built model: validate in Molprobity, and
then rebuild (Coot obviously ...).
.... while doing this check the following which just takes computer
time, not yours:
-twining: try any de-twining tool, I would simply switch on twining
refinement in REFMAC and sit back and read the log carefully.
-wrong space group: Try the Zanuda server: http://www.ysbl.york.ac.uk/YSBLPrograms/index.jsp

regards -

Tassos

(*)on the other hand I did bet on Germany winning against Spain, so my
betting skills are worse than these
of a cephalopod mollusk named 'Paul', http://en.wikipedia.org/wiki/Paul_the_Octopus

PS When you use TLS the average B will always go down since a lot of
the movement is 'absorbed' by the TLS tensors
that describe domain movement - what is in the PDB is just the
'residual' B that cannot be explained by domains moves.

On Jul 8, 2010, at 8:32, Tim Gruene wrote:

> Dear Sampath,
>
> You are right, the gap between R and Rfree is significant and
> indicates that
> your model was overfitted.
> Without knowing your data or your model, some reasons for
> overfitting might be:
> - you used automated placement of water molecules (e.g. through
> arpwaters or in
> coot) and never checked the water molecules for chemical
> reasonability. How
> many residues are there in your structure and how many water
> molecules?
> - there might be a domain that - despite the resolution - does not
> resolve with
> your data but you built somethig nevertheless
> - you build things into your model while using a too low (approx.
> <1.0sigma)
> sigma-level for your map. At too low a contour level you can often
> see what
> you _want_ to see in my experience, and not what is there
> - you screwed up the Rfree set and it's not indendent anymore.
> However, in that
> case I would rather expect the difference or the ration to be too
> small rather
> than too big.
> - Your data may be twinned.
>
> That's just a first set of reasons but there might be something one
> could only
> know by looking at your data.
>
> Tim
>
>
>
> On Wed, Jul 07, 2010 at 10:17:14PM -0700, Sampath Natarajan wrote:
>> Dear all,
>>
>> I have a question about the R free value. I refined a structure
>> with 2A
>> resolution. After model building and restraint refinement using
>> Refmac
>> program, the average B factor was around 50 for all atoms. The R/
>> Rfree were
>> around 22/34. Then used the TLS refinement choosing entire
>> molecule. Then
>> R/Rfree reduced as 20/32. But the average B factor was reduced as
>> 30. The
>> R/Rfree difference is about 12% in final refinement. I feel it is
>> significantly higher.
>>
>> Could any one suggest me to reduce the Rfree value more? or is it
>> good to
>> submit the data in the PDB database with this 12% difference?
>>
>> Thanks for the suggestions.
>>
>> Sincerely,
>> Sampath N
>
> --
> --
> Tim Gruene
> Institut fuer anorganische Chemie
> Tammannstr. 4
> D-37077 Goettingen
>
> GPG Key ID = A46BEE1A
>

P please don't print this e-mail unless you really need to
Anastassis (Tassos) Perrakis, Principal Investigator / Staff Member
Department of Biochemistry (B8)
Netherlands Cancer Institute,
Dept. B8, 1066 CX Amsterdam, The Netherlands
Tel: +31 20 512 1951 Fax: +31 20 512 1954 Mobile / SMS: +31 6 28 597791

CCP4bb navigation

CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

Previous message:
Subject: Re: Question about R/Rfree value difference
From: Tim Gruene tg {- at -} SHELX {- dot -} UNI-AC {- dot -} GWDG {- dot -} DE
Date: 2010-07-08

Next message:
Subject: Re: Question about R/Rfree value difference
From: Tim Gruene tg {- at -} SHELX {- dot -} UNI-AC {- dot -} GWDG {- dot -} DE
Date: 2010-07-08