Re: [ccp4bb] Refmac5 twin refinement pushing Rfree surprisingly down

Try MR with one copy in all space groups of PG 321/312 using Phaser. Going from PG 3 to PG 32 should halve the number of copies per ASU. You may have to re-process your data in the higher point group to do this. Or you might actually have a tetartohedral twin, but just try with the higher-symme

Re: [ccp4bb] Refmac5 twin refinement pushing Rfree surprisingly down

Hi Keller, Thanks for the suggestions! I only have two copies in ASU at SG P32. Zanuda also suggests P32 is the best SG. On Thu, Apr 13, 2017 at 8:12 PM, Keller, Jacob wrote: > Yes, this was my case exactly—it looks like there are two pairs of coupled > twin domains: a,c and b,d. Assuming you h

Re: [ccp4bb] Refmac5 twin refinement pushing Rfree surprisingly down

Yes, this was my case exactly—it looks like there are two pairs of coupled twin domains: a,c and b,d. Assuming you have multiple copies of your model in the same ASU, try doing MR in higher symmetry space groups of point group 312 or 321, like P3212 etc. There is this handy page with all the spa

Re: [ccp4bb] Refmac5 twin refinement pushing Rfree surprisingly down

Hi Keller, I do not how to check twin fraction after Refmac (I guess it's somewhere in log file). From the log file it seems I have four twin domain: Twin operators with estimated twin fractions Twin operator: H, K, L: Fraction = 0.275; Equivalent operators: K, -H-K, L; -H-K, H,

Re: [ccp4bb] Refmac5 twin refinement pushing Rfree surprisingly down

What was the refined twin fraction after Refmac? It’s much more accurate than initial tests. Also, how many twin domains do you have? If you have many, it might be a higher space group but with less twinning. I recently had a case in which apparent tetartohedral (four-domain) twinning in P32 was

Re: [ccp4bb] Refmac5 twin refinement pushing Rfree surprisingly down

Twin refinement cannot be compared directly to untwinned - the R factors are between different parameters - without twinning it is assumed you have an amplitude obtained more or less from sqrt(I But for a twinned data set that I is actually [ I1 + twin_factor I2 ] so the amplitude is not really c

Re: [ccp4bb] Refmac5 twin refinement pushing Rfree surprisingly down

Hi Alex, You are not giving the number after refinement without the twin refinement. Nevertheless, R-free drops like this are not unheard of. You should check your Refmac log file, it would warn you of potential space group errors. Refmac will also give you a refined estimate of the twin fract

[ccp4bb] Refmac5 twin refinement pushing Rfree surprisingly down

Dear All, I have a protein/dna complex crystal and data collected at 3A and another set at 2.8A, space group P32. L test shows twinning (fraction around 0.11). The structure solved by MR and model building of the complex finish (no solvent built yet, I do not think it's good to build solvent in su

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Dear Colleagues, The deadline for application to the annual CCP4/APS school in macromolecular crystallography: “From data collection to structure refinement and beyond” is in less than a week - April 19. Please read the thread below for basic information on the school. For all details, please vi

Re: [ccp4bb] in crystallo enzymatic activity

Are you using the same columns for purifying both WT and mutant? Could you have the tiniest fraction of WT contaminating your mutant batches which would then turnover you substrate in crystallo over the couple of weeks? You could try washing the columns/systems with NaOH or pepsin to remove WT,

[ccp4bb] in crystallo enzymatic activity

Dear CCP4bb, We work on an enzyme that we crystallized with two substrates bound in the active site (the reaction transform two substrates into two products). We have also the structure with the two products. We are able to see densities for the substrates when we collect data at differen

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Re: [ccp4bb] waters with positive FoFc peaks?

How are the peaks coordinated? Regular coordination with 5 or more ligands has ion written all over it. Different difference density peak heights can be caused by differences in effective B-factor restraint weight but also by differences in relative scattering factors of oxygen and whatever you

Re: [ccp4bb] waters with positive FoFc peaks?

It would be useful to know what wavelengths you were talking about. Also, try an anomalous difference Fourier map to see whether the atoms are weakly anomalous. From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Andrew Marshall Sent: Thursday, April 13, 2017 2:00 AM To: CCP4B

Re: [ccp4bb] waters with positive FoFc peaks?

Partially occupied Cs+ comes to mind. At lower resolution the difference may be more easily fudged away by the refinement lowering the B. Mark J van RaaijCNB-CSICwwwuser.csic.es/~mjvanraaij Original message From: Andrew Marshall Date: 13/04/2017 07:59 (GMT+01:00) To: CCP4BB@J