Hi All,
The problem has also been solved with a new 2.0A dataset collected
over the last weekend. Same space group and dimensions, much less
radiation damage. This time I used APS SER-CAT's weaker BM beamline.
Thanks,
Tongqing
Tongqing Zhou, Ph.D.
Staff Scientist
Structural Biology Section
Vaccine Research Center, NIAID/NIH
Building 40, Room 4607B
40 Convent Drive, MSC3027
Bethesda, MD 20892
(301) 594-8710 (Tel)
(301) 793-0794 (Cell)
(301) 480-2658 (Fax)
******************************************************************
The information in this e-mail and any of its attachments is
confidential and may contain sensitive information. It should not be
used by anyone who is not the original intended recipient. If you
have received this e-mail in error please inform the sender and
delete it from your mailbox or any other storage devices. National
Institute of Allergy and Infectious Diseases shall not accept
liability for any statements made that are sender's own and not
expressly made on behalf of the NIAID by one of its representatives.
******************************************************************
-----Original Message-----
From: Zhou, Tongqing (NIH/VRC) [E]
Sent: Tuesday, June 15, 2010 10:45 AM
To: [email protected]
Subject: Re: [ccp4bb] Stuck refinement
Hi, Everyone,
Thank you all very much for the nice suggestions. I am trying to
reply within this email.
I agree that the problem may be rooted from the crystal itself, we
noticed during data collection that a wedge of the rotation was very
mosaic, HKL2000 was able to pick up the right spots, but the scaling
gives high chi^2, and when I used the rejection files, HKL2000
complained "more than 50000 rejections". Colleagues suggested
tweaking the error model, the complaint of "more than 50000
rejections' went away and rejection dropped to below 300 spots. The
new error model reduced the chi^2 as well as the I/sigI in the low
resolution shells.
I run the P222 data set with Xtriage, the report says no twining,
but the symmetry was too low. However, HKL2000 won't even pick up
higher symmetry groups during indexing. I also rescaled the data
omitting the bad wedge, xtriage gives "normal" report.
Refinement was done with combination of simulated annealing, TLS,
ADP, individual sites in Phenix. The molecular replacement was done
with CDR-loop-trimmed antibody Fab and antigen structures. The map
quality was good and I was able to rebuild the new loops without any
problem.
I will have beam time later this week, I think it will be better to
put a better crystal on.
Best regards,
Tongqing
Tongqing Zhou, Ph.D.
Staff Scientist
Structural Biology Section
Vaccine Research Center, NIAID/NIH
Building 40, Room 4607B
40 Convent Drive, MSC3027
Bethesda, MD 20892
(301) 594-8710 (Tel)
(301) 793-0794 (Cell)
(301) 480-2658 (Fax)
******************************************************************
The information in this e-mail and any of its attachments is
confidential and may contain sensitive information. It should not be
used by anyone who is not the original intended recipient. If you
have received this e-mail in error please inform the sender and
delete it from your mailbox or any other storage devices. National
Institute of Allergy and Infectious Diseases shall not accept
liability for any statements made that are sender's own and not
expressly made on behalf of the NIAID by one of its representatives.
******************************************************************
-----Original Message-----
From: Eleanor Dodson [mailto:[email protected]]
Sent: Tuesday, June 15, 2010 4:46 AM
To: Zhou, Tongqing (NIH/VRC) [E]
Cc: [email protected]
Subject: Re: [ccp4bb] Stuck refinement
When this happens, I firstly suspect that the spacegroup may be
wrong. We had a case where the symmetry was pseudo I4212 but was
really
I222 (or was it really I212121) Anyway most of the structure obeyed
the
I41212 symmetry but there was a tail which did not..)
Feed the unmerged reflections into pointless and see what it suggests
Eleanor
Zhou, Tongqing (NIH/VRC) [E] wrote:
Hi Everyone,
I have some problem in refining a structure. The data goes to 2.4A
(with some 30% completeness at 2.15A), the structure was solved by
MR with Phaser, refinement was done with Phenix, but the r and r-
free are now staying at 26% and 32%, even with all possible waters
and missing fragments added. Data was collected at APS at cryo
condition. One thing I noticed during HKL2000 data processing was
that the chi^2 were way too high at lower resolutions shells, I
had to adjust the default error model in HKL2000 to get the chi^2
to around 1, but this adjustment reduced the overall I/sigI ratio a
lot (from around 20 to 5).
The quality of electron density maps looks fine to me for a 2.4 A
data set and I was able to build all the missing CDR loops for the
antibody in the complex. I am lost now, should I just re-collect a
new data set?
Thanks,
Tongqing
Tongqing Zhou, Ph.D.
Staff Scientist
Structural Biology Section
Vaccine Research Center, NIAID/NIH
Building 40, Room 4607B
40 Convent Drive, MSC3027
Bethesda, MD 20892
(301) 594-8710 (Tel)
(301) 793-0794 (Cell)
(301) 480-2658 (Fax)
******************************************************************
The information in this e-mail and any of its attachments is
confidential and may contain sensitive information. It should not
be used by anyone who is not the original intended recipient. If
you have received this e-mail in error please inform the sender and
delete it from your mailbox or any other storage devices. National
Institute of Allergy and Infectious Diseases shall not accept
liability for any statements made that are sender's own and not
expressly made on behalf of the NIAID by one of its representatives.
******************************************************************
