Hi Francisco,
you could try using these keywords in Mosflm:
POSTREF WIDTH 3
#since you have 0.1 degree oscillation and Mosflm can only handle 30
images at once, the default I believe is 5 images if I'm not mistaken
SEPARATION CLOSE
PROFILE RMSBG 25
POSTREF USEBEAM
Then I would start indexing from various images separated let say by 15
degrees use 6 or more if you have, don't go to full resolution first
refine the cell estimate the mosaicity, fix the mosaicity and process a
low res pass (what is your resolution ?).
I assume you didn't run strategy beforehand and checked where you would
have overlaps ? If you still have the crystal you could collect another
dataset in a different orientation perhaps (assuming you can move
kappa). I would then index on two images run strategy and get some
images where the predicted overlaps are, then try to optimize the
distance to the detector in favour of no overlaps. If you can, use the
biggest CCD detector available or image plate if that has a larger
detector area. Since you're in P222 you could also offset 2theta to
artificially increase your detector area. Set the beam position almost
at the edge of the detector and give your crystal a good 360 or more
degree spin - sure you will end up with somewhere close to 100GB of data
but who cares if you can solve the problem. 650 A is a challenge but not
impossible - how many molecules in the asu do you expect ? Can you solve
your problem with MR or do you need HA phases - if that is the case,
then searching for another crystallization condition might be faster.
Another thing to mention is if you have a homehexamer don't forget you
can use NCS averaging so if you "only" collect a 2.5 A dataset your
resulting electrondensity maps might look like 2 A after averaging. I
suggest to rather be modest in resolution and have no overlaps then be
able to solve the structure and go back eventually to optimize for
higher resolution etc.
With the cell dimensions you obtained you could startup XDS if you want
to give it a try. As Graeme mentioned xia2 might be easier for a first
time XDS user.
Regarding obtaining new / different crystals you could try the Hampton
additive screens with your crystal condition, with some luck you might
end up with a more favourable crystal form and smaller cell dimensions.
Assuming you are working on a macromolecular complex, do you have any EM
information perhaps - then you would know if e.g. your cell dimensions
correspond to one complex perhaps or more. Then the smallest cell axis
you will most likely get will be in the size of your complex.
Just some thoughts,
Juergen
[EMAIL PROTECTED] wrote:
Dear All
I am processing data from a crystal for a large macromolecular complex
with mosflm. Cell dimensions are around 120 x 150 x 650, with a p222
spacegroup. To avoid overlaps, we have collected data with a oscillation
of 0.1 degrees.
When I try to process the data with mosflm, mosaicity decreases along the
processing to a very low values (0.05 to 0.1 depending on the images). The
result is that I miss a lot of spots from the images.
I would appreciate any help regarding :
1.- What's is the best strategy to process such a dataset ?
2.- Which are the critical parameters in mosflm to avoid these problems,
and how to modify them ?
3.- and finally, can I use this data or your advice is to try to get
crystals in a different spacegroup ?
Many thanks
Best regards
Francisco
-----------------------------------------
Francisco J. Enguita, Ph.D.
Macromolecular Crystallography Laboratory
ITQB
EAN, Av. da República
2781-901 Oeiras
Portugal
Phone : +351-21-4469663
Fax : +351-21-4433644
E-mail : [EMAIL PROTECTED]
-----------------------------------------
--
Jürgen Bosch
University of Washington
Dept. of Biochemistry, K-426
1705 NE Pacific Street
Seattle, WA 98195
Box 357742
Phone: +1-206-616-4510
FAX: +1-206-685-7002
Web: http://faculty.washington.edu/jbosch
