The problem was resolved by Isabel Uson and Giovanna Petrillo using
ARCIMBOLDO!
The real spacegroup is P21, twinned to look like C2221. I would
probably take years to solve that on my own, they did it very fast. This
was a difficult problem, and still is, since if I try MR using the
correct solution and space group I cannot resolve it. I definitely
recommend everybody with difficulties to give Arcimboldo a try. Thank
you Isabel!
Thank you Sergei Strelkov, your message was very informative as
usual. ;)
Thank you George M. Sheldrick for the information on how to
discrimitate between correct and wrong MR solutions using SHELXE.
Best regards,
Napo
On 11/4/2011 7:42 AM, Sergei Strelkov wrote:
Dear Napoleão,
Thank you for updating everyone on your
efforts, and also acknowledging the advice.
I wanted to respond to your question regarding maps.
I know that many people who try to figure out
whether or not their MR solution is the right one
would ask the same question.
So first of all if you wonder why you actually get very
decently looking maps the answer is a classical one:
because 'the phases are more important than amplitudes'.
The appearance of your map is defined by
your model phases, and hence a good match between the
model and the map /may not/ be taken as
a sign of a correct solution. Once again: /never ever!/
On the contrary, at least in your coil1.jpg
image I clearly see that the density exactly follows
the model which is almost entirely poly-Ala.
Unless your protein is really poly-Ala this should be
alarming. If you had a correct solution they you
would hope to see the (difference)density for
at least some missing side chains.
And a second point. Unless your model contains
the complete chain (which is rarely the case, especially
for the coiled coils, as discussed already)
a sign of the correct solution would be the appearance
of extra density near the N- and/or C-terminus of
the model. If it is not there, it is almost certainly not
a solution.
And you should not be worried about the R-factors being
very high at this stage. If the solution is correct then
you should see at least some extra features in the map.
Kind regards,
Sergei
Thank you all for the replies.
Sorry for taking so long to reply, I was actually trying some of your
interesting ideas (and I'm still trying).
I tried using the low resolution data sets for the molecular replacement
(thanks to Yuriy Patskovsky), I also improved and increased my coiled
coil database and employed it in many approaches using EPRM (interesting
program I was not aware of), which I found to produce lots of data,
hopefully addressing at some extent the helixes bent (thanks to Bernhard
Rupp). I also tried some more tweaking in Phaser, although not sure if
did it properly (thanks to Randy Read).
There is no twinning as far as I can tell (thanks to Ed Pozharski for
the tip). Using a data set with enough completeness (360 degrees @
Brookhaven) and processing in P1 did not help me because in this space
group there is most likely 2-3 helixes in the asymmetric unit, which
complicates the problem (and it takes a lot of time for Phaser to run).
Automated approaches also did not yield a better result (as far as I can
tell). I'm convinced that the space group is C2221, but I may be wrong.
Thanks to Sergei Strelkov for the numerous useful suggestions on how to
approach the problem.
One of the big issues for me is to discriminate between a lot of
similarly good density maps. For example:
http://www.fullonline.org/coils/coil1.jpg
http://www.fullonline.org/coils/coil2.jpg
I have hundreds of solutions like these and I think they are all wrong.
I couldn't manage to run Arcimboldo, could not find a tutorial on it
either. It was highly recommended here (and elsewhere), so I'm
definitely willing to give it a try (thanks Isabel Uson).
You guys opened my eyes about a series of issues that I should learn
about and approach, I'm most thankful for that.
Best regards,
Napo
--
Prof. Sergei V. Strelkov
Laboratory for Biocrystallography
Department of Pharmaceutical Sciences, Katholieke Universiteit Leuven
O&N2, Campus Gasthuisberg, Herestraat 49 bus 822, 3000 Leuven, Belgium
Work phone: +32 16 330845 Fax: +32 16 323469 OR +32 16 323460
Mobile: +32 486 294132
Lab pages:http://pharm.kuleuven.be/anafar
