From: Giorgio Giardina
Date: 8 February 2012 11:41
Hello,
I have some interesting small molecule xtals.
I was wondering if it is possible to collect a small molecule data-set using a sincrotron macromolecular xtallography beam line, maybe with a very low beam intensity and moving the detector as close as possible?
Has anybody experienced that?
And if I get the images back home, can I process them using standard macromolecular software or do I need ab-initio special programs?
Will MR work for phasing?
Thanks in advance,
Giorgio
----------
From: Tim Gruene
Hello Giorgio,
most synchrotron beamlines should have a resolution limit beyond or near
1A resolution which is sufficient for solving the structure with direct
methods.
At least XDS has no problems with non-chiral space groups and can be
used to process the data. Since with a small cell you are going to have
only few spots per image, make sure you increase the DELPHI parameter to
30 or 60, as Kay Diederichs pointed out to me.
XDS_ASCII.HKL can be read into xprep which, if you keep on hitting enter
and provide it with the chemical composition when it asks you to, is
going to prepare a shelxd input file that can be used to solving the
structure with shelxd. Its output .res-file is the starting point for
refining the structure with e.g. shelxl.
So: yes, you can process the data with [sparkle ;-)] standard
macromolecular software [/sparkle]
Cheers,
Tim
- --
- --
----------
From: Kevin Jin <kevinjin@gmail.com>
Date: 8 February 2012 16:02
To: CCP4BB@jiscmail.ac.uk
I collected GTP/Mg2+ crystal on SSRL beamline 9-1 before. The images
was processed by Mosflm and structure was solved by Shelx as usual.
Kevin
----------
From: Martin Walsh
Hi Giorgio, there are beamlines dedicated to small molecule crystallography at synchrotrons as well. I can suggest I19 at Diamond (obviously) but there are others!
http://www.diamond.ac.uk/Home/Beamlines/I19.html
Martin
----------
From: Jens Kaiser
Giorgo,
We have done that routinely for quite some time now. We had problems
when using a normal CCD detector, where we had to collect two or three
sweeps to avoid overloads (see below). Since we have the PILATUS this is
not necessary anymore and the data behaves fine. "Problems" still
persisting are: we have only a single axis goniometer, which can lead to
low completeness in P1 and P-1. Highest energy (17keV) and closest
distance (188mm) at our beamline have many SM crystals (even the ones
that "don't diffract" in house -- that is a 300 or 500 u sealed tube)
with an I/sig of 5-10 at the edge of the detector. Crunch, Acorn,
ShelxCDE and ShelxS don't have any problem with any of the data we
collected to <0.9A resolution. The multipass caused some inexplicable
non definite positives during refinement. We haven't tracked that down
systematically, so it might just have happened haphazardly.
HTH,
Jens
----------
From: Santarsiero, Bernard D.
Most beamlines have attenuators, so there's little reason to collect
multiple sweeps. We always collect 360deg. Since it's a small molecule,
and usually fairly large and robust, you can warm it up, nudge it in a
different direction with a pin (we use sterile, disposable acupunture
needle), and refreeze it in the cryostream. Then do a second sweep in a
different orientation.
I recommend moving the beam energy to 15.5KeV or higher to compress the
diffraction image. Collect with 5-10deg widths. We can typically get the
detector to around 70-80mm. You need to get to 0.85A resolution or better
for good, stable refinement, and Acta Cryst. requires that resolution for
publication. Often you need the low-resolution data and data to better
than 1A to help with the sigma2 relationships in direct methods.
You see both primary and secondary extinction, and that extinction can be
anisotropic, so the SWAT option in SHELX is most useful. Otherwise, the
overall scale factor is off, typically overestimated by the strong
low-resolution reflection intensities, with the result that the
anisotropic Gaussian displacement parameters may become non-positive
definate.
Bernie
--
Bernard D. Santarsiero
----------
From: Kay Diederichs
Hi Giorgio,
some XDS-related hints can be found at
http://strucbio.biologie.uni-konstanz.de/xdswiki/index.php/Small_molecules
which I renamed to "Difficult datasets" since some of the suggestions also apply to those.
What is lacking in that article is that you really should specify SENSOR_THICKNESS= and SILICON= . This is already taken care of in the Pilatus XDS.INP templates,
but for CCD detectors this has to be specified manually; see hints in the script http://strucbio.biologie.uni-konstanz.de/xdswiki/index.php/Generate_XDS.INP
HTH,
Kay
----------
From: Gordon Leonard
Hi Giorgo,
Just to say that we routinely, and often, do these sorts of experiments on ID29 and ID23-1 at the ESRF using an energy of 20 keV (~0.62 Angstrom wavelength) and detector distances that allow collection of data to a resolution of better than 0.7A. Mini-kappa goniometers also allow for the collection of data in more than one crystal orientation in the case of completeness problems.
As has already been pointed out by others who have replied to your posting, the processing of the images with MOSFLM or XDS is usually pretty straightforward as is structure solution using direct methods.
Hope this helps
Gordon
----------
From: Giorgio Giardina
You have been all wonderfully helpful.
The landscape is crystal-clear now.
Thanks to everybody.
Giorgio
Date: 8 February 2012 11:41
Hello,
I have some interesting small molecule xtals.
I was wondering if it is possible to collect a small molecule data-set using a sincrotron macromolecular xtallography beam line, maybe with a very low beam intensity and moving the detector as close as possible?
Has anybody experienced that?
And if I get the images back home, can I process them using standard macromolecular software or do I need ab-initio special programs?
Will MR work for phasing?
Thanks in advance,
Giorgio
----------
From: Tim Gruene
Hello Giorgio,
most synchrotron beamlines should have a resolution limit beyond or near
1A resolution which is sufficient for solving the structure with direct
methods.
At least XDS has no problems with non-chiral space groups and can be
used to process the data. Since with a small cell you are going to have
only few spots per image, make sure you increase the DELPHI parameter to
30 or 60, as Kay Diederichs pointed out to me.
XDS_ASCII.HKL can be read into xprep which, if you keep on hitting enter
and provide it with the chemical composition when it asks you to, is
going to prepare a shelxd input file that can be used to solving the
structure with shelxd. Its output .res-file is the starting point for
refining the structure with e.g. shelxl.
So: yes, you can process the data with [sparkle ;-)] standard
macromolecular software [/sparkle]
Cheers,
Tim
- --
- --
----------
From: Kevin Jin <kevinjin@gmail.com>
Date: 8 February 2012 16:02
To: CCP4BB@jiscmail.ac.uk
I collected GTP/Mg2+ crystal on SSRL beamline 9-1 before. The images
was processed by Mosflm and structure was solved by Shelx as usual.
Kevin
----------
From: Martin Walsh
Hi Giorgio, there are beamlines dedicated to small molecule crystallography at synchrotrons as well. I can suggest I19 at Diamond (obviously) but there are others!
http://www.diamond.ac.uk/Home/Beamlines/I19.html
Martin
----------
From: Jens Kaiser
Giorgo,
We have done that routinely for quite some time now. We had problems
when using a normal CCD detector, where we had to collect two or three
sweeps to avoid overloads (see below). Since we have the PILATUS this is
not necessary anymore and the data behaves fine. "Problems" still
persisting are: we have only a single axis goniometer, which can lead to
low completeness in P1 and P-1. Highest energy (17keV) and closest
distance (188mm) at our beamline have many SM crystals (even the ones
that "don't diffract" in house -- that is a 300 or 500 u sealed tube)
with an I/sig of 5-10 at the edge of the detector. Crunch, Acorn,
ShelxCDE and ShelxS don't have any problem with any of the data we
collected to <0.9A resolution. The multipass caused some inexplicable
non definite positives during refinement. We haven't tracked that down
systematically, so it might just have happened haphazardly.
HTH,
Jens
----------
From: Santarsiero, Bernard D.
Most beamlines have attenuators, so there's little reason to collect
multiple sweeps. We always collect 360deg. Since it's a small molecule,
and usually fairly large and robust, you can warm it up, nudge it in a
different direction with a pin (we use sterile, disposable acupunture
needle), and refreeze it in the cryostream. Then do a second sweep in a
different orientation.
I recommend moving the beam energy to 15.5KeV or higher to compress the
diffraction image. Collect with 5-10deg widths. We can typically get the
detector to around 70-80mm. You need to get to 0.85A resolution or better
for good, stable refinement, and Acta Cryst. requires that resolution for
publication. Often you need the low-resolution data and data to better
than 1A to help with the sigma2 relationships in direct methods.
You see both primary and secondary extinction, and that extinction can be
anisotropic, so the SWAT option in SHELX is most useful. Otherwise, the
overall scale factor is off, typically overestimated by the strong
low-resolution reflection intensities, with the result that the
anisotropic Gaussian displacement parameters may become non-positive
definate.
Bernie
--
Bernard D. Santarsiero
----------
From: Kay Diederichs
Hi Giorgio,
some XDS-related hints can be found at
http://strucbio.biologie.uni-konstanz.de/xdswiki/index.php/Small_molecules
which I renamed to "Difficult datasets" since some of the suggestions also apply to those.
What is lacking in that article is that you really should specify SENSOR_THICKNESS= and SILICON= . This is already taken care of in the Pilatus XDS.INP templates,
but for CCD detectors this has to be specified manually; see hints in the script http://strucbio.biologie.uni-konstanz.de/xdswiki/index.php/Generate_XDS.INP
HTH,
Kay
----------
From: Gordon Leonard
Hi Giorgo,
Just to say that we routinely, and often, do these sorts of experiments on ID29 and ID23-1 at the ESRF using an energy of 20 keV (~0.62 Angstrom wavelength) and detector distances that allow collection of data to a resolution of better than 0.7A. Mini-kappa goniometers also allow for the collection of data in more than one crystal orientation in the case of completeness problems.
As has already been pointed out by others who have replied to your posting, the processing of the images with MOSFLM or XDS is usually pretty straightforward as is structure solution using direct methods.
Hope this helps
Gordon
----------
From: Giorgio Giardina
You have been all wonderfully helpful.
The landscape is crystal-clear now.
Thanks to everybody.
Giorgio
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