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Recently I had a similar situation for some very long, thin needles.
Radiation damage occurs for crystals of both the wild type and
selenomethionyl-substituted proteins, and the rate of decay of the
diffraction pattern in the x-ray beam is virtually indistinguishable
for the two types of crystals if they are of comparable thickness. In
order to obtain a reasonably redundant SAD dataset to 2.9 angstroms
resolution, I simply collected small, overlapping wedges of data (about
50 degrees per wedge, but each wedge started at 25 degree intervals) at
various translations along the crystal. Since the Laue symmetry was
orthorhombic P222 this meant I only needed 4 wedges of data. Upon
scaling the data in HKL2000_ph I realized that I only needed to use 35
degrees in each data wedge to achieve a complete, redundant (6-fold)
dataset that was suitable for structure solution. It was apparent from
the scaling statistics that the diffraction in the last 15 degrees of
each data wedge had decayed considerably, and the inclusion of this
data degraded both the overall diffraction limit and the anomalous
signal from the selenium atoms. A similar strategy was used to collect
a wild type dataset (on thicker crystals) to a diffraction limit of
2.55 angstroms (vs. 2.85 angstroms for a standard data collection
strategy, without crystal translation).
This data was collected at the APS on the Structural Biology Center's
19-ID beamline, where the size of the beam at the crystals was
approximately 120 x 120 microns. This worked well for crystals that on
average are about 30 x 500 microns in size.
If your crystals aren't long enough for you to take advantage of such a
data collection strategy, then why not try collecting data on multiple
crystals and merging the data from multiple crystals into one dataset?
That's how we all used to collect data back in the dinosaur-age before
cryocooling at near liquid nitrogen temperatures.
Diana
Dear bulletin members,
Sorry for non-CCP4 related questions.
Recently, I've got crystals which have a radiation damage problem.
It also has a weak diffraction because of the big unit cell dimension
(about 300A in one direction), so I have to overexpose crystals even
with
the synchrotron beam. The problem is that during the data collection,
the
resolution decaded gradually from 3 to 6A after certain number of
frames
(about 30 or 40 frames). Do you have any experiences or any suggestions
that I can evade this radiation damage? Any suggestions will be
grateful.
Thank you.
Best regards,
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
Uhn-Soo,Cho
Graduate student in Biological Structure Department
University of Washington.
HSB G514
1959 NE Pacific St.
Seattle, WA 98195-7420
Box. 357420
Fax #206-543-1524
Tel # 206-221-2435
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
* * * * * * * * * * * * * * * * * * * * * * * * * * * *
Diana R. Tomchick
Associate Professor
University of Texas Southwestern Medical Center
Department of Biochemistry
5323 Harry Hines Blvd.
Rm. ND10.214B
Dallas, TX 75390-8816, U.S.A.
Email: [EMAIL PROTECTED]
214-645-6383 (phone)
214-645-6353 (fax)
