findncs
[Keyworded input]
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F I N D N C S
Detecting NCS relations from heavy atoms sites
Version 1.1 Sep-30-1997
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The program requires at least 6 heavy atom sites for each NCS one operations unit i.e. at least 3 sites for each NCS asymmetric unit (which can be a protein monomer, dimer, trimer or even higher oligomer). Once the coordinates of the sites were input, they were extended by crystallographic symmetry and lattice repetition. Then the program systematically searches whether a group of sites can match another group of sites by an NCS operation. There are a number of criteria to choose one NCS operation among crystallographic symmetric related ones:
In some cases, for example, when the number of sites is low enough (less
than 20), the space group is simple (such as triclinic, monoclinic, R3, P3),
and/or the NCS relations are regular (like 222 tetramer 2-fold dimer and
so on) users can run the program fully automatically and accept the suggestions
from the output. However, if users have high symmetry, many HA sites or
many protein molecules with non-regular NCS relations, they have to
follow the instructions in this manual to find the NCS step by step.
Alternatively, users can also define the search range (which includes
the entire oligomer of the protein) themselves using knowledge from the
solvent flattening map. Otherwise, it might take long CPU time and or
the results are too complicated to understand. For details, please read
through Key Word Command and Frequently Asked Questions of this document.
Examples
CELL, COMPOUND, DISP, ERROR, FRCLIM, FSITE, LIST, MAXNCS, MINMATCH, SITE, SPACEGROUP, SPHERE, XYZLIM
example: fsite 0.755 0.282 0.146 or fsite 0.689 0.467 0.299 PCMBif the compound name is not given, the program will assume the name is the same as the default name from COMPOUND. CELL command must be given before this command.
examples site -13.752 16.271 27.267if the compound name is not given, the program will assume the name is the same as the default name from COMPOUND. See convention of the orthogonal system. If the coordinates are already put into the correct asymmetric unit, user does not have to give cell dimension and space group.
example -1. 1. -1. 1. -1. 1.By default, the program extends HA sites to 8 unit cells (XYZ from -1. to 1.), to make sure that the search range includes the entire protein molecule. However, if users have many crystallographic operations, (like cubic, hexagonal or tetragonal), it is extremely time consuming to find out in such a range and there are a lot of chances to have "noise" (false NCS). So, users have to determine a smaller search range themselves (see FAQ 4.a). This command can be repeated. See also XYZLIM and SPHEre
example -10 50 -100 40 -20 60
This command can be repeated; see FRCLIM
The program uses following convention for orthogonal system
X is in A direction
Y is in AB plane
Z is perpendicular to AB plane
Each NCS operation can be described as that an object
rotate kappa degree about a certain axis, then move a
screwing distance along the direction of this axis
In the following for each possible NCS, the program provides:
Matching pairs: number of matching pairs by the possible NCS solutions
is ranked by this number
Matching members: the ID number of each site (from input order before
crystallographic operation)
RMS: root mean square deviation of the superimposed sites
Screw: the screwing distance
Radii: average distance between center and all joining site
Polar angles: they describes the NCS axis as follow
psi is the angle between Z and NCS axis
phi is the angle between X and image of NCS axis XY plane
kappa is the rotation angle. 180=2fold, 90=4-old ... so on
the polar angle definition is same as in the POLARRFN program
Center: center position of all the joining sites, this
can help to find out if two axis interact each other
Example
Maximum number of NCS operations to be output 10
Space Group >>> P21 4
Symmetric operation ---- Total: 2 Rotation: 2
8 sites were read in
16 sites after symmetry operations
116 sites are extended to maximum cells
Building a distance matrix......
Looking for NCS matches......
Total 285 NCS operations have been found
Maximum atom number 6
generating unit cell frame for O...
----------------------------------------------------------------------
NCS1 with matching pairs 6
1 2 3 4 5 7
4 3 2 1 7 5
NCS matrix:
-0.99584 0.08831 0.02261
0.08831 0.87302 0.47961
0.02261 0.47961 -0.87719
65.24355 -19.55399 64.35049
RMS: 0.929 Screw: 0.00 Radii: 41.20
Polar angle: 75.65 87.30 180.00 & 104.35 -92.70 -180.00
Center: 32.07 -21.44 29.19
----------------------------------------------------------------------
NCS2 with matching pairs 6
2 3 4 5 7 8
3 2 8 7 5 4
NCS matrix:
-0.98548 -0.15782 -0.06262
-0.15782 0.71543 0.68063
-0.06262 0.68063 -0.72995
-39.70554 -30.00050 66.40594
RMS: 0.993 Screw: 0.00 Radii: 54.58
Polar angle: 111.56 -84.74 180.00 & 68.44 95.26 -180.00
Center: -21.63 4.36 40.89
----------------------------------------------------------------------
NCS3 with matching pairs 6
2 3 4 5 6 7
3 2 6 7 4 5
NCS matrix:
-0.99408 0.10813 0.01057
0.10813 0.97522 0.19301
0.01057 0.19301 -0.98114
67.17181 -9.37817 58.34059
RMS: 1.197 Screw: 0.00 Radii: 40.59
Polar angle: 84.43 86.87 180.00 & 95.57 -93.13 -180.00
Center: 32.69 -21.11 27.57
----------------------------------------------------------------------
NCS4 with matching pairs 5
2 4 6 7 8
4 2 7 6 3
NCS matrix:
0.88325 0.04691 0.46656
-0.05447 -0.97798 0.20146
0.46574 -0.20335 -0.86124
-38.24167 57.20192 92.85443
RMS: 0.948 Screw: -14.65 Radii: 36.55
Polar angle: 75.94 -0.12 167.96 & 104.06 179.88 -167.96
Center: -22.16 21.90 50.91
----------------------------------------------------------------------
NCS5 with matching pairs 5
1 2 4 6 8
8 4 2 7 1
NCS matrix:
-0.99553 0.01697 -0.09289
0.01986 -0.92409 -0.38165
-0.09232 -0.38179 0.91963
51.18448 -21.65644 -2.32794
RMS: 1.292 Screw: 0.48 Radii: 56.42
Polar angle: 168.44 76.36 179.92 & 11.56 -103.64 -179.92
Center: 25.93 -9.40 -8.48
.....
CPU time: 1 min 20.5 sec
2) Can I directly use the NCS matrix from FINDNCS in averaging programs?
It depends on the circumstance of protein oligomers. For example,
if you only have a dimer AB in crystallography asymmetric unit (asu), the
program can easily find the NCS from A to B (or B to A). However, if you have
tetramer ABCD, the program will give you the NCS matrix A_to_B, A_to_C,
A_to_D, B_to_C, B_to_D and C_to_D, with total 6 matrices while in
most averaging programs you probably only need first 3 matrix. However, your
tetramer is in 222 symmetry, A_to_B/D_to_C should be the same matrix, so
is A_to_C/B_to_D, A_to_D/B_to_C. The program in this case will only
output 3 matrix which you can use in the averaging program.
Sometimes the program does not output the NCS which you want, for example biologically, ABCD is the tetramer biologically, the program can give NCS A_to_B and A'_to_F' where A' and F' are crystallographically equivalent to A and F. You can find it when you try to make the mask before averaging. You can use SPHERE and XYZLIM or FRCLIM.
Although FINDNCS does not always give you the exact NCS operation required by averaging programs, it is still much much faster to find out the correct NCS using the program as a tool than find NCS by hand.
2.a) How should I analyse the output of FINDNCS?
Look at the graphics using the outpdb files and O files.
>From the log output.
For example, you find the following output shows a dimer of 2-fold
symmetry. The joining sites 1-fit-4 and 2-fit-3.... after the 180 deg NCS
operation. (only in two fold symmetry, site 1 fit 4 and while 4 also fit 1
in the same NCS)
----------------------------------------------------------------------
NCS1 with matching pairs 12
1 2 3 4 5 6 7 8 9 10 11 12
4 3 2 1 8 7 6 5 12 11 10 9
NCS matrix:
-1.00000 -0.00061 0.00025
-0.00061 0.70840 -0.70581
0.00025 -0.70581 -0.70840
0.74407 -0.21214 -0.51413
RMS: 0.091 Screw: 0.00 Radii: 30.48
Polar angle: 67.55 -89.98 180.00 & 112.45 90.02 -180.00
Center: 0.37 -0.01 -0.30
If you find the following NCS in the same time,
----------------------------------------------------------------------
NCS2 with matching pairs 12
1 2 3 4 5 6 7 8 9 10 11 12
2 1 4 3 6 5 8 7 10 9 12 11
NCS matrix:
....
RMS: 0.210 Screw: 0.00 Radii: 30.48
Polar angle: 157.55 -89.46 180.00 & 22.45 90.54 -180.00
Center: 0.37 -0.01 -0.30
----------------------------------------------------------------------
NCS3 with matching pairs 12
1 2 3 4 5 6 7 8 9 10 11 12
3 4 1 2 7 8 5 6 11 12 9 10
NCS matrix:
....
RMS: 0.210 Screw: 0.00 Radii: 30.48
Polar angle: 89.82 0.10 180.00 & 90.18 -179.90 -180.00
Center: 0.37 -0.01 -0.30
>From the matching site number, you can find this is a perfect 222 NCS
symmetry only from the output. Of course it will be much easier to understand
if you look at graphics using the PDB files (and O files if you use O).
3) How to display the results by graphics?
If the DISPLAY command is present, the program will generate some files for graphics display. First, it generates a PDB file ncsall.pdb which include all the sites within the search range. After NCSs have been found, the program will generate ncs1.pdb ncs2.pdb ..... which includes sites joining operations of NCS1, NCS2 ... and so on. You can use any graphics program to display them.
In the case you use O and/or RAVE:
The program also generates a file called ncs.ofm which include NCS
operators, and vectors which can be used by the O program. Then program
will make an O macro file called oncs.mac (together with ncs1.mac, ncs2.mac...).
If users run O under the same directory and type @oncs.mac, there will be
a group of commands appearing in menu bar (@ncs1.mac, @ncs2.mac). If
you want to display first NCS operations found by FINDNCS, click
@ncs1.mac the O program will display a axis and the sites which join the
NCS in yellow and sites superimposed by this NCS.
After running @oncs.mac The NCS matrix is stored as
.lsq_rt_ncs1
.lsq_rt_ncs2
....
If you have bones, you can display as two objects (e.g. SKEL and SKEL1)
If you want to see how the 1st NCS works type:
lsq_obj ncs1 SKEL to see how it is superimpose to SKEL1
If you have a electron density call map1, you can use command:
lsq_rt_obj .lsq_rt_ncs1 map1 to superimpose this map.
If you edit the file oncs.mac to take out .lsq_rt_ncs1... to a new file,
it can be directly used by the RAVE packages.
4) What should I do in the case the program takes an intolerable CPU time?
The time of the program is proportional to N*N*N*N*(N-1), where N
is the total number of search sites after crystallographic operations. If
this number is more than 300, the calculation will become very slow (10 hours
in a DEC-alpha for 310 atoms, so about 300 hours for 620 atoms). If users
use an automatic mode for XYZ limit (8 unit cells XYZ from -1 to 1), this
number can be found in the following line
736 sites are extended to maximum cells
In this case users have to use smaller search range defined by commands
FRCLIM, XYZLIM or SPHERE. (The sites number inside selected range can be
found in the line:
148 sites are inside the selection ranges )
Increasing MINMATCH is also the way to substantially
decrease the CPU time when too many possible solutions are found. If the
program output is something like this
Solution: 1000 Max match: 16
i1,i2,i3,j1,j2,j3 1 83 91 57 115 67 CPU time: 103.0 s
Solution: 2000 Max match: 16
i1,i2,i3,j1,j2,j3 4 47 76 58 36 85 CPU time: 379.2 s
You find out that the maximum matching number is 16 so setting minmatch to 6
should not hurt anything.
4.a) How should I define the search range?
The idea of defining a search range is to include at least one entire
protein oligomer so the program would not miss the correct NCS. If a search
range includes 8 crystallographic ASU homogeneously in XYZ directions, I think
(not proved) an entire protein oligomer won't be missed
in more than 95% cases.
It does not matter if the search range includes more than one entire protein oligomer. (The program should be able to recognize the crystallographic symmetric equivalent NCSs by finding the joining sites of these NCSs are crystallographic equivalent too.) However, too big a search range might cause many sites to be searched and slow down the search. It is not very practical if the sites inside the range are more than 300.
There are several ways you can decide the search range.
MATCH # 3 with matching pairs 5 MATCH # 4 with matching pairs 5 MATCH # 15 with matching pairs 5 ........ MATCH # 1585 with matching pairs 12 MATCH # 1569 with matching pairs 13 MATCH # 681 with matching pairs 14 MATCH # 1070 with matching pairs 14Now you know the best match so far is MATCH 681. You open the file findncs.log and find MATCH 681 and you can find NCS there. The atom file ncsall.pdb has been generated and you can use graphics to see what this NCS looks like and choose the search range round the joining sites.