Direct method solution at 1.15A

From: Yuri Pompeu
Date: 23 September 2011 19:49

Hello everyone,
I have a data set >99% completeness to 1.15A
This is a 400 amino acid long protein and it has 7 Met (Sulfur peaks around 20sigma)
And a tightly bound phosphate (P peak around 22sigma)
Could I try and solve this directly or is it crazy idea?
If so what program should I try?

thanks
Yuri


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From: George T. DeTitta


Yes you should. AND you are crazy. One of George's SHEL programs or perhaps SnB from the Hauptman group. Good luck!

Sent via BlackBerry by AT&T

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From: Jacob Keller

I don't really get your question: assuming the sigmas you mentioned
are in an anomalous map and therefore have been located, why don't you
just plug it into your usual phasing algorithm?

Jacob

--
*******************************************
Jacob Pearson Keller
Northwestern University
Medical Scientist Training Program
*******************************************

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From: Yuri Pompeu

I solved the structure using molecular replacement. Those sigmas are simply from my sigmaa 2mFo-DFc maps.
I was wondering if I could try and solve sort of like small molecules are....

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From: George M. Sheldrick

Dear Yuri,

There is a general rule that you need data to 1.2 Angstroms or better to
solve a structure by ab initio direct methods. To be more precise, more
than half the reflections between 1.1 and 1.2 A should have I>2sigma(I).
So you are probably just within this limit. However it appears that the
largest structures solved this way with no atom heavier than S were not
more than half the size of your protein. The programs most often used
for such crazy attempts are SnB, SIR and SHELXD. It would be a good idea
to find a workstation with at least 32 CPUs and run the multi-CPU version
of SHELXD and be patient, it might take a few weeks.

If you just want to remove model bias (a serious problem with MR
solutions using 3A data, but no problem with 1.15A data) you could try
MRSAD phasing. There are many good programs for doing this, but a
particularly simple way would be to use ANODE to find the anomalous
sites from your MR solution and the new beta-test SHELXE for the
density modification and tracing. The only other program you would
need for this is SHELXC. I am still developing these programs but am
happy to provide them on email request. No other programs, libraries
etc. are needed because these programs have zero dependencies.

Best wishes, George

--
Prof. George M. Sheldrick FRS
Dept. Structural Chemistry,
University of Goettingen,
Tammannstr. 4,
D37077 Goettingen, Germany


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From: Fan, Hai-fu

Dear Yuri,

If you have located all the heavy atoms (sulfur and phosphor) correctly, you could try sulfur-SAD phasing using the program OASIS. This program has a record of solving a 1206 residues protein with SAD signals from 22 sulfur atoms scattered under Cu-Ka radiation and a record of solving a 213 residues protein with SAD signals from 2 sulfur atoms scattered under Cr-Ka radiation.

By the way please note that the OASIS in CCP4 6.2.x is not the uptodate version. You can get a better version from http://cryst.iphy.ac.cn. The latest version will be available on the website in the coming October.

 

Best regards,

Hai-fu

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From: Eleanor Dodson


At this resolution you may/ should be able to find anamalous scatterers using the anom signal from P and S. the SHELXC/D/(E) package is very good at this!

Once you have the sites for the heavier atoms I would expect most direct methods programs couild  extend that sub-structure.
Certainly ACORN seems to work without resolution limits, and give excellent final maps.

Eleanor