Hi Kostya, phenix.resolution reports effective resolution of data set. It is based on methods described here: J. Appl. Cryst. (2015). 48, 589-597 Acta Cryst. (2013). D69, 1921-1934 As I pointed out before, a single number for highest resolution isn't very informative unless you show data completeness per resolution (in phenix.refine log file it is reported in the beginning of macro-cycles right after bulk-solvent step). Pavel On 10/7/16 00:15, Kogan, Konstantin wrote:

Dear all,

Thank you for your comments. The maps are not that bad, and actually from biological point of view we do see everything we want. One of the questions is how to report the parameters for the structure, as I have a feeling that the effective resolution is much lower than 2.64Å, which is the current limit, and the mean B value is twice higher than estimated Wilson B value. All together it seems that this is the data I have, but I would like to be sure, that I have got maximum out of it.

I attach the xtriage log file for more information, if needed. Also I attached the example of a good map region at 1.6 sigma and a bad map region at 1.0 sigma.

I have also tested to refine at 2.8, 2.9, and 3.0, and I haven't seen much difference in maps quality, but the Rwork/Rfree went down a bit.

Isn't is seems a bit suspicious 2.64Å resolution, Rwork/Rfree=0.28/0.31, and mean B value 114? Is there a way to present/report effective resolution?

Kostya

Hi Kostya,

how complete the data set is? Missing reflections in some resolution zones between 2.64A and inf may be sufficient to make the effective resolution lower or much lower than 2.64A which would explain why the map does not look like what you expect.

Pavel

On 05/10/16 22:53, Randy Read wrote:

...

Hi,

As Christian said, the level of anisotropy is not that bad for this structure. In terms of what Phenix can do to handle it, first, when you solved it by MR, Phaser would have accounted explicitly for the anisotropy. In fact, Phaser’s algorithms underlie the anisotropy server that produced your plot! Phenix.refine will also refine the overall anisotropy parameters.

When you have really severe anisotropy, the current algorithms may not account well enough for the differing accuracy of reflections in the weak directions. So it can sometimes be helpful to do anisotropic pruning of the data in the way that the anisotropy server implements it. But this is something to do with some caution, and I think you should only press on with it if the maps allow you to see things you couldn’t see clearly before.

Best wishes,

Randy Read

----- Randy J. Read Department of Haematology, University of Cambridge Cambridge Institute for Medical Research Tel: +44 1223 336500 Wellcome Trust/MRC Building Fax: +44 1223 336827 Hills Road E-mail: [email protected] Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk

...

On 5 Oct 2016, at 20:32, Christian Roth wrote:

Hi Kostya,

I have seen worse data than that, which are ususally treated quite well within phenix using standard parameters. With a resolution of about 2.7 Ang. I wouldn't expect to see waters and also just well ordered side chains, whereas more flexible ones just give some "bumps" pointing away from the main chain density. Though without a picture or something else it is difficult to say if your maps are really unusually bad.

Cheers

Christian

Am 05.10.2016 um 07:47 schrieb Kogan, Konstantin:

...

Dear all,

I have highly anisotropic data. I was able to solve the structure by MR, and refine it with phenix.refine to Rwork/Rfree=0.28/0.31 with Resolution cutoff 2.64Å and mean B values 114, which is pretty high for such resolution. Though the overall structure makes sense, the maps are quite featureless, no water molecules, and side-chains refinement is a problem. I have tested how the data is anisotropic with Diffraction Anisotropy Server (see attached image), which shows clearly, that we don't really have data to 2.64Å in all directions. Though, there are other servers/programs that can deal specifically with anisotropic data e.g. STARANISO anisotropy server and then refine with BUSTER, I would like to know if it's possible still to use Phenix with some more advance parameters to actually address the issue of anisotropy and to get maximum out of the data. The space group is P61 2 2, cell parameters are 73, 73, 453, 90, 90, 120 and I have high multiplicity data, but no NCS.

Thanks in advance for any input,

Kostya -- Konstantin (Kostya) Kogan Postdoctoral researcher Pekka Lappalainen's Lab Institute of Biotechnology University of Helsinki Helsinki, Finland Mobile: +358-(0)45-8994342

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