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It would be very nice if you could confirm the presence of NCS by a self
rotation function.
Be aware of the fact that the bromides are/might be very susceptible to
radiation damage. Signs of specific damage can be seen from data
processing statistics (chi-square vs frame) in favorable cases.
( see figure 1 or ref 4 (below) )
If the redundancy of the data allows it, you could chop the data set in
2 halves and use isomorphous differences in substructure location and
phasing. You could also use unmerged data in SHARP to model the
disapearance of the Bromide atoms (if that is happening.)
Some (biased) set of relevant references:
1. Ennifar, E., Carpentier, P., Ferrer, J.-L., Walter, P. & Dumas, P.
(2002). Acta Cryst. D58, 1262-1268.
(Difficulties with MAD data due to radiolytic debromination.)
2. Ravelli, R. B. G. & McSweeney, S. M. (2000). Structure, 8, 315-328.
(Rip phasing.)
3. Schiltz, M., Dumas, P., Ennifar, E., Flensburg, C., Paciorek, W.,
Vonrhein, C. & Bricogne, G. (2004). Acta Cryst. D60, 1024-1031.
(using unmerged data in phasing while taking care of disapearing atoms)
4. Zwart et al, Acta Cryst. (2004). D60, 1958-1963 ( using both ano and
rip signal in the location and phasing in a rather naive but effective
way.)
HTH
Peter
William Scott wrote:
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Hi folks:
I have a question regarding a Br mad data set on RNA.
We have a crystal of an RNA in the lab in which the molecule has been
transcribed and incorporates 11 Br-U residues at known positions in the
sequence, and we have a MAD data set on this to about 3 A resolution (as
well as a native data set).
mmtbx.xtriage reports that we may have up to 3 molecules in the
asymmetric unit (of P21), meaning 11, 22 and 33 Br sites are possible:
----------------------------------------------------------------
| Copies | Solvent content | Matthews Coef. | P(solvent cont.) |
|--------|-----------------|----------------|------------------|
| 1 | 0.828 | 7.146 | 0.012 |
| 2 | 0.656 | 3.573 | 0.238 |
| 3 | 0.484 | 2.382 | 0.652 |
| 4 | 0.312 | 1.786 | 0.091 |
| 5 | 0.140 | 1.429 | 0.007 |
----------------------------------------------------------------
| Best guess : 3 copies in the asu |
----------------------------------------------------------------
I've been using ShelxD to find Br sites, and the maps, though quite
promising, still need significant improvement. I also haven't convinced
myself that there is a clear NCS 2-fold or 3-fold, and so have been
trying with 1,2, and 3 molecules.
Since there are regions of this molecule that (presumably) have
canonical A-form RNA helices, and these have Br-U in them at known
positions, I'd like to make use of the distance constraints these put on
the Br sites, as well as the Br absorption in the mad peak data.
Is it possible to do molecular replacement using the Br anomalous signal
and model helices to help pin down the locations of the Br atoms?
Thanks in advance.
Bill Scott
