DM for skeletonisation (CCP4: General)
NAME
dm_skeletonisation
- iterative skeletonisation using dm
This document describes the iterative skeletonisation process as
implemented in 'dm', and in particular the things you need to know in
order to use the process.
THE SKELETONISATION PROCESS
The process of skeletonisation involves two stages:
-
The construction of a 'skeleton' representing the connectivity of the
map.
-
The creation of a new map from the skeleton.
These stages are conducted as follows:
1. Construction of the Skeleton
First, the map is searched to locate the important features. These
fall into two classes: Peaks, and Join points.
Peaks are local maxima in the density. A ridge is the highest path
joining two peaks. A join point is the lowest point on a ridge,
i.e. the lowest point on the highest path joining two peaks.
In constructing the skeleton, there are two important parameters to
consider:
-
The join point cutoff. Any ridge whose join point falls above this
level will be included in the skeleton. Ridges whose join points fall
beneath this level will not be included in full, but a part of the
ridge may be included as an end.
-
The end point cutoff. If a join point is below the join point cutoff,
a portion of the ridge may still be added to the skeleton if the peak
is above the end point cutoff. That portion of the ridge which is
above this level is added to the skeleton.
Obviously, ridges included under (1) will contribute to the
connectivity of the map, whereas ridges included under (2) will not
because they are always dead-ends. Thus it is normal to set the end
point cutoff somewhat higher than the join point cutoff in order to
enhance connectivity in the map.
To simplify the setting of these parameters, 'dm' determines the
cutoff levels by examining the map in order to produce a skeleton with
particular features. These features are specified as the length of
skeleton (per residue) which should be generated from join point
ridges, and the length of skeleton (per residue) which should be
generated from dead-end ridges. It is my hope that these optimum
values of these parameters should be fairly constant, and so that for
most cases the default values should suffice. If new values are
required, they can be set using the SKEL LENGTH card.
One other parameter affects the skeletonisation process: the
temperature factor applied to the initial map. Skeletonisation based
on the sharpened map used by 'dm' is fairly poor, a much better
skeleton is obtained if the map is smoothed with an overall temperature
factor first. Again this parameter is not very sensitive and can be
left at the default value, you may possibly want to increase it if the
initial map is particularly poor or decrease it if the initial map is
very good.
2. Map Generation
Map generation is carried out by building up a cylindrically symmetric
density distribution about the skeleton. The distribution is taken
from an atom averaged over a cylinder whose length is the average
protein atomic bond length.
Density outside the solvent mask is set to the mean solvent value.
USING SKELETONISATION
In skeletonisation calculations I have found the best scheme is to
perform a conventional density modification first, and then use the
modified map as a starting point for a dm/skeletonisation calculation.
The phase extension scheme 'SCHEME AUTO FRAC 0.5' is a good option for
this second calculation, as reflections with high figures-of-merit are
more likely to be included in the starting set.
Note also that it is wise to make sure the final cycle of the
skeletonisation run is a conventional density modification cycle rather
than a skeletonisation cycle otherwise your building may become
excessively biased towards the 'dm' skeleton. Thus for 'SKEL EVERY 2'
you would set 'NCYC' to be odd, for 'SKEL EVERY 3' (the default) you
would set 'NCYC' to be (3n-1) where n is an integer.
A typical command file is given below:
dm
hklin ~/hkl/gmtomir.mtz
hklout $SCRATCH/gmt_dm.mtz
<< 'MY-DATA'
SOLC 0.35
MODE SOLV HIST
NCYC 10
SCHEME RES FROM 3.7
LABI FP=FP SIGFP=SIGFP PHIO=PHIB FOMO=FOM
LABO PHIDM=PHIdm FOMDM=FOMdm
END
'MY-DATA'
#
dm
hklin $SCRATCH/gmt_dm.mtz
hklout $SCRATCH/gmt_sk.mtz
<< 'MY-DATA'
SOLC 0.35
MODE SOLV HIST SKEL
SKEL EVERY 2
NCYC 9
SCHEME AUTO FRAC 0.5
LABI FP=FP SIGFP=SIGFP PHIO=PHIB FOMO=FOM PHIDM=PHIdm FOMDM=FOMdm
LABO PHIDM=PHIsk FOMDM=FOMsk
END
'MY-DATA'
AUTHOR
Dr. Kevin Cowtan, Protein Structure Group, University of York, England
|