Re: [ccp4bb] Twin questions: is my crystals twinned or not?

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

Previous message:
Subject: Twin questions: is my crystals twinned or not?
From: Parthasarathy Sampathkumar spartha2 {- at -} GMAIL {- dot -} COM
Date: 2010-07-08

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Subject: Re: Twin questions: is my crystals twinned or not?
From: Ethan Merritt merritt {- at -} U {- dot -} WASHINGTON {- dot -} EDU
Date: 2010-07-08

Subject: Re: Twin questions: is my crystals twinned or not?
From: Anastassis Perrakis a {- dot -} perrakis {- at -} NKI {- dot -} NL
Date: 2010-07-08

Hi Partha -

A few thoughts:

1. If you attach logs, at least gzip them ...
2. From the fact that Rfree goes down when twinning is switched off, I
would think there is no twining.
3. The intensity distribution (moment of E, etc) suggest no twining.
The twining operator you use is the crystallographic two fold, I think.
4. During refinement the relative fraction of twin is a refined
parameter, to make you model+twining fit the data. The initial twin
fraction is only an estimate.
5. Did you consider P3121 or P3221 at all? I think its unlikely, but
easy to check in Phaser as alternative space group.
6. Do these R/Rfree include TLS refinement and waters? if not they are
quite Ok, given the missing residues
7. I see you only have 430 free reflections. I would argue they are
too few to get a reliable Rfree and would aim for more, 10% instead of
5% will give you about 900. Be careful though in not choosing a new
Rfree for the refinement from now on.

Good luck -

A.

On 8 Jul 2010, at 20:03, Parthasarathy Sampathkumar wrote:

> Dear All,
>
> Back ground:
> This is my first experience with a twined dataset. Crystals belong
> to a small domain of 132 aa, out of which ~40 residues appears to be
> disordered (~30 of those from C-terminal and C-term His6 tag).
>
> Initial space group: P3 with unit cell dimensions: 62.507 62.507
> 55.117 90.00 90.00 120.00; Resolution 2.35Angs.
>
> Pointless suggested P321 (space group # 150) as possibility. I
> determined structure by MR with Phaser (1 molecule in ASU). After
> several model building and refinement cycles R and Rfree got stuck
> at 24.0% and 31.6% respectively.
>
> Therefore, I considered P3 space group (now Two molecules in the
> ASU) with a twin component. I was only able to add handful residues
> to the model already refined in P321. My current R and Rfree factors
> are 21.0% and 29.1%, respectively for Two molecules refined in P3
> space group.
>
> Questions:
>
> 1. H-test in cTruncate suggested a twin fraction of 0.43 for the
> twin operator -h-k, k, -l. Where as Refmac5 with Amplitude based
> twin refinement gave an initial value of 0.508 for the same
> operator. Why these values are different between cTruncate and
> Refmac5 (is this because I asked Refmac5 do amplitude Twin
> refinement instead of Intensity based)?
>
> 2. I noticed in Refmac5 log file that twin fractions changes for
> every cycle of refinement. During my most recent Refmac5 run Initial
> estimate of 0.508 for -h-k, k, -l operator changed to 0.504 at the
> end of 20th cycle. The corresponding values were 0.519 and 0.529,
> respectively, in a previous round. Since twin estimates were based
> on measured Intensities (in turn amplitudes) why would they change
> with refinement (am I missing something here)?
>
> 3. When I repeated my final round of Refmac5 WithOut Twin Refinement
> my R and R-free factors are 22.9% and 28.6%, respectively, which
> also appears to be OK for 2.35 Angs. data (in fact, slightly better
> R-free). These values are likely to improve little bit after
> completing the solvent model. So, is this crystal really twinned?
>
> I have attached log files of cTruncate and most recent Refmac5 run
> with Twin refinement. Apologies for attachments (at least no image
> files).
>
> Thank you all in advance for educating me.
>
> -Partha
>
>
>
> <34_truncate_anl.log><40_refmac5.log>

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

Previous message:
Subject: Twin questions: is my crystals twinned or not?
From: Parthasarathy Sampathkumar spartha2 {- at -} GMAIL {- dot -} COM
Date: 2010-07-08

Next message:
Subject: Re: Twin questions: is my crystals twinned or not?
From: Ethan Merritt merritt {- at -} U {- dot -} WASHINGTON {- dot -} EDU
Date: 2010-07-08