Hi Alphar,
A while ago I had success with a poor MIR map by building in a few
fragments I could identify and then combining phases from this model
with the MIR phases using SIGMAA (CCP4 program) and then calculating
new maps. A couple of cycles of this resulted in phases in which I had
confidence and I was able to build the complete structure. (There's
probably a more modern way to do this now ...)
Cheers,
Charlie
Quoting Fengyun Ni <[email protected]>:
Thanks, Ezra!
I have a native dataset.
I'll try what you said, Hope it works in my case.
Thanks!
Alphar
2009/2/21 Ezra Peisach <[email protected]>
Personally, I would try to build into what you see and then do
phase
recombination. I had a SeMet crystal in which I could only see
part of a
sheet and helix after DM... Things were not connected - until I
improved
the phases with my model. Also - do you have a native dataset?
You do not
mention it.
Ezra
Fengyun Ni wrote:
Hi everyone!
I have a question on my poor MIR map. Four datasets (two AU and
two HG)
were used for phasing upto 3 A resolution in my case. I could
locate several
heavy atom sites for each dataset with occupancy finally refined
to about
0.3 to 0.4, and with B value refined to about 50 to 80. I did not
include
the anomalous data because once I refine against anomalous data,
the
anomalous occupancy was only about 0.02, and even smaller as
0.004 for some
sites. I guess the anomalous data are not good enough, so I did
not use them
right now (Could I do this? Or must I include them somehow?).
The FOM I got is about 0.6, the Cullis-R factors were about 0.6
for all
four data sets, and the phasing power was about 2.5 in each
dataset. After I
did the density modification, the FOM could increas to about 0.8.
My problem is that the protein should form a long helix
structure as
indicated by other homology protein, but in the map after density
modification, the densities are not consecutive though the
overall shape
seemed to be a long helix. The good thing is that some short
helix-like
densities could be observed in the current map, but they are not
connected
to each other.
Right now, I am totally lost whether I should believe in the
mir-map I
have. Could I improve this map with some other method? Or should
I
re-do the
phasing?
BTW, the space group for my crystal is R3 or R32 with a=50, b=50,
c=380,
alpha=90, beta=90 and gamma=120. The c-axis is much longer than
a- and
b-axis, so the reflection data suffered from the anisotroy
effect.
Any suggestion on my phasing problem is welcome.
--
Alphar Ni
Call me alphar~
:)
--
Fengyun Ni
Call me alphar~
:)
--
Charlie Bond
Professorial Fellow
University of Western Australia
School of Biomedical, Biomolecular and Chemical Sciences
M310, 35 Stirling Highway
Crawley WA 6009, Australia
