Dear Matt,
as Eleanor pointed out your obtained solution is certainly correct, and for
2.8ang resolution almost as good as it could get.
But could it not just be that the residual density you observed in your
electron density ("There is some density for the receptor ") isn't the
receptor but another Fab fragment? Speaking from own experiences that is
very often the case if you are working on antibody-antigen complexes.
Cheers
Stefan
-----Ursprüngliche Nachricht-----
Von: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] Im Auftrag von
Eleanor Dodson
Gesendet: Donnerstag, 23. Juli 2009 14:50
An: CCP4BB@JISCMAIL.AC.UK
Betreff: Re: [ccp4bb] is my crystal twinned or not?
It seems very likely your crystal is twinned - the moments and
cumulative intensity are a good indicator of twinning, especially when
there isnt much NCS.
But your solution is almost certainly partially correct - the Rfactor is
pretty low.
It is a good idea to run pointless which gives you the correlation
Coefficient for each symmetry operator. see if there is any difference
between the different ones.)
You could have twinning with either 4-fold or 222 symmetry.
the 222 is less likely seeing that would mean the a=b axes are
accidently equal.
If you decide the symmetry is really PG 4 you need to remerge the data
in that point group ( no need to reintegrate - the cell dimensions will
still be the same)
The most likely solution is that one of your P43212 molecules is
corrrect, but it is probably a good idea to run the MR search again.
Use your refined antibody molecule as the search model to save work later..
Eleanor
Matthew Franklin wrote:
> Hi all -
>
> I'm trying to solve the structure of an antibody-receptor complex, and
I've hit a wall which may be due to a twinned crystal form. This crystal
has the apparent space group P43212, with cell constants a=64.02 c=274.83.
Solvent content analysis using this space group suggests 1 mol/asu, with 47%
solvent, which would be fairly consistent with the diffraction limit of
about 2.8 A. I've been able to place the antibody using molecular
replacement and refine it to R=0.289, Rf=0.338. There is some density for
the receptor, which represents about 20% of the mass of the complex, but not
clear enough to build into, and all efforts to improve the density or place
the receptor by molecular replacement have failed.
>
> Well, tough luck, you might say, but I noticed at the very beginning of
the process that this crystal form may be a perfect twin. The "4th moment
of E" graph from Truncate shows nearly all resolution bins with values of
1.4 - 1.6, except in the very topmost few bins where the values rise up to
2. The other moment graphs are likewise at their "perfect twin" values
across the resolution range. The cumulative intensity distribution graph
shows the observed values are about 30% lower than the expected values for
both acentric and centric reflections. As I understand it, both of these
are strong indicators of twinning, and the twin fraction analysis in DETWIN
suggests a twin fraction of ~0.5.
>
> So what do I do now? I think I'm supposed to reprocess the data in the
space group without the twin transformation (which would be P43), then run
molecular replacement which should show me solutions for both halves of the
twin. However, all I'm seeing is two copies of the antibody in the (P43)
asymmetric unit, which don't overlap, and both of which need to be present
in the same unit cell in order to form a 3-dimensional lattice. This is
exactly what I would expect to see if there were no twinning present.
>
> So my question is, is this crystal form twinned or not? Is there some way
that the intensity statistics could be misleading me? On the other side, am
I doing something wrong with the structure determination if the crystal is
twinned? How should I proceed?
>
> Thanks for any help anyone can provide.
>
> - Matt
>
>
> --
> Matthew Franklin , Ph.D.
> Senior Scientist, ImClone Systems,
> a wholly owned subsidiary of Eli Lilly & Company
> 180 Varick Street, 6th floor
> New York, NY 10014
> phone:(917)606-4116 fax:(212)645-2054
>
>
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