Anomalous signal for chlorides

From: Yuri Pompeu
Date: 25 November 2011 23:31


Hello everyone,
I have a good data set to 1.17A that I solved by MR.
I come accross some sites that appear to be chlorides. I was
wondering if they could have some anomalous signal.
I also have a well ordered phosphate  and a couple of S from Met´s.
How do I go about probing the signal from these?
Thank you. Best,

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From: Boaz Shaanan

Hi,

You could calculate an anomalous difference map using your current phases and see whether you see any peaks around the atoms you suspect are Cl, S or P (although the latter should have a clear tetrahedral geometry, certainly at your resolution). It all depends of course on the wavelength at which you collected your data (check Ethan Merritt's site), but again at your resolution you may very well hit lucky.

  Cheers,

             Boaz


Boaz Shaanan, Ph.D.

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From: Yuri Pompeu


Hi Boaz,
Yes indeed, the phosphate group of the molecule looks quite beautiful at 1.17A and it has a really big peak 18sigma!
Is there a utility for calculating anomalous maps, or is it simply an option in the refinement program?

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From: Pavel Afonine


Hi Yuri,

phenix.refine, for example, always outputs Fourier map coefficients for anomalous difference map if input input data contain Fobs+ and Fobs- (or Iobs+ and Iobs-).

Pavel

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From: George Sheldrick


Dear Yuri,

The new program ANODE (J.Appl.Cryst. (2011) 44, 1285-7, Open Access) is designed for this and is very simple to use. It may be downloaded from the SHELX beta-test server (please email me if you require downloading intructions).

George

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From: Randy J. Read

Dear Yuri,

In addition to the other good options that have been presented, you can use the log-likelihood gradient maps in Phaser to find anomalous scatterers. We find this to be very sensitive, and it has the advantage of being iterative (i.e. when you find some anomalous scatterers, this improves your model and thus the sensitivity for finding additional sites).

When you're starting from a refined model, we think it is best to look for purely imaginary scatterers to add to the real scattering in your model. In the ccp4i GUI, choose the "SAD with molecular replacement partial structure" mode, provide the data (with F+ and F-), the wavelength, and the current model, then turn on LLG-map completion and select the "Complete with purely anomalous scatterer" option.

If you want to see the initial LLG map, set the number of completion cycles to 0 and turn on the option to output log-likelihood-gradient map coefficients. Open these in coot as a difference map, choosing the columns FLLG_AX and PHLLG_AX. If you let Phaser complete the sites, then the final LLG map should be nearly flat and you have to look at the PDB file containing the sites that it found.

Best wishes,

Randy Read

--
Randy J. Read

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From: Jacob Keller


Not that Phaser needs my approval, but I recently did exactly what
Randy recommended and it really found basically all of the S and Cl
sites, with data at resolution 2.2 Ang and wavelength 0.979 Ang, too.
I also played a bit with the sigma cutoff for adding new sites so that
the stronger sites (Se in my case) are found but the weaker ones not.
Also, don't forget to click the "output LLG map coefficients" option
to get the right columns in your mtz.

Jacob

--
*******************************************
Jacob Pearson Keller
*******************************************

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From: Eleanor Dodson


There are many ways of course to do this..
But if you are using the CCP4 GUI, refmac refinement it is a 2 step process.

CAD to add the DANO SIGDANO columns from the input data back into the refmac output

fft
 Select anomalous map

Use DANO SIGDANO PHIC FOM asinput

Switch on the peak search

Read that pdb into coot and check that the highest peaks are on anomalous scatterers - you usually find the S are visible too..

 If the data is reasonable I do it routinely to verify results - are the SO4s really sulfates etc etc..

Eleanor