On 21/09/2020 14:49, Qiuye Li wrote:
Yes, I extract one single subunit just to quickly access the shape or
the sequence of the ordered region; next I would like to model 3~5
subunits.
The reason I think segmentation is necessary is, many subunits at the
ends of the filament were half chopped off (by masking in relion). So,
to have a map consisting of entirely full subunits, I tried to clean
up the ends. However, even doing so ended up replicate display
problems in Coot (just tested a "clean" 5mer in a 125*109*59 box). It
seems that as long as I use segment->select->save selected region
function in Segger, it will cause such problems. So I guess the
question now is, how should one extract a subregion of a map without
causing display problems in Coot?
1) It is not yet clear to me that there is any problem with Coot.
2) I am unconvinced that extraction of a sub-region is needed. If I
wanted to see the shape of the density or side chains, I'd look down the
z axis and narrow down the clipping.
3) If there are sub-units at the ends of the filament that are chopped
by masking, then build in a region where the masking hasn't chopped the
map. I don't think that you need a map consisting entirely of full
sub-units. You just need a few - maybe only one.
4) If this is like the tau filaments built by my colleagues over the
last couple of years it should be straight-forward, shove in a few beta
strands, link them and extend them as needed... add the symmetry-related
copies as I mentioned yesterday. The only non-trivial problems (IIRC)
were the sequence assignment and wondering if the strand went up or down
for the next rung at the corner with bad/missing density.
Paul.
Any information/background/suggestion are welcome!
Best,
Qiuye
On Sun, Sep 20, 2020 at 10:41 PM Paul Emsley
<[email protected] <mailto:[email protected]>> wrote:
On 20/09/2020 19:27, Qiuye Li wrote:
The protein is a filament and the reconstructed map is in a
512*512*512 box. I segmented this reconstructed map in Chimera
using Segger, selected a single asymmetrical unit, and saved it
as a new map (which was referred to as the segmented map). This
new map automatically changed to a box size of 94*84*26.
That does seem a bit thin in z. Does segger know about helical
symmetry? If not, I don't think that this is a winning strategy.
But it does occur to me to wonder why you even need to segment the
map. Relion tells you the helical symmetry and you should (it
seems to me) just apply that to the model.
if you even need to do that... I would build a model into the set
of tubes in the middle, and then when I wanted to be concerned
about symmetry, just try cut paste rotate translate (just shove it
up a rung and use jiggle fit) and let the NCS chain copy tool
modify the other chains as I go. From what I can see of your map,
one chain either side of the central chain should be enough.
Paul.
BTW, I tried to display from another direction and it looks like
these copies do lie along the axis, see the attachment.
Best,
Qiuye
On Sun, Sep 20, 2020 at 1:29 PM Paul Emsley
<[email protected] <mailto:[email protected]>> wrote:
On 20/09/2020 17:29, Qiuye Li wrote:
>
> Thanks for the suggestion, it looks map segmentation and
coot do not
> work well with each other, see the tests I did below:
>
> 1. .mrc map directly from relion:
> in Chimera: OK
> in Coot: OK
>
> 2. segmented map generated from Chimera:
what do you mean by "segmented"?
> in Chimera: OK
> in Coot: displayed as replicates
>
> When I segment the map, the box dimensions and map
threshold level
> were changed, would these cause problems?
>
I am not really sure what you mean by the map threshold
level. It's
something of a surprise that the box (cell) dimensions have
changed.
Maybe that's a result of reboxing? But I doubt that that
would cause
problems to Coot.
It looks like the map is interspersed with sections that have
zero
value. However, from the orientation axes top left it doesn't
seem that
the sections lie along the direction of a cell axis.
Paul.
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