GETAX (CCP4: Supported Program)

NAME

GETAX - real space correlation search

PURPOSE

Real space searching for rotation axis of a D<n> or C<n> multimer ( <n> = 2,3,4,5,6,... ).

If you have:

a multimer in the asymmetric unit

some initial phases

a peak in a selfrotation function

you can start using this program to find the translational part of your NCS operators.

It has worked in several cases with even very poor phases ( 20 molecules/au, <fom>=0.25 to about 6Å resolution).

VERSION

Version 2.5 (14. April 1998)

SYNOPSIS

getax MAPIN foo.map [ XYZIN foo1.pdb ] [ XYZOUT foo2.pdb ] [ MAPOUT bar.map ]
[Keyworded input]

DESCRIPTION

GETAX is a program to search for your non-crystallographic symmetry if a first map is available. The only knowledge you need is a selfrotation solution (from e.g. POLARRFN) and a crude knowledge of the size/shape of your molecule(s).

INPUT/OUTPUT FILES

MAPIN

map covering a whole unit cell with axis order X=fast, Y=medium, Z=slow changing index.

This map can be a 6 Angstroem MIR map on a 2 Angstroem grid. So if you use fft you don't have to worry about getting the grid right, since fft takes 1/3rd of the high resolution anyway. Sometimes it can be helpful to try finer grid spacings (this slows down the calculation, though).

Depending on your spacegroup, you will have to change the extent and/or axis order with MAPMASK.

XYZIN

a PDB file with an initial model for the correlation search.

see: INPUT XYZ

XYZOUT

a PDB file with the initial search sphere/slice as build and used by GETAX.

This can be a PDB file with orthogonal coordinates either before (OUTPUT XYZ) or after initial interpolation (OUTPUT GXYZ).

To be sure that everything works fine: have a look at this output file with your favourite graphics program (O or RASMOL or whatever ...): there should be no overlap between the segments and the rotation axis should be properly oriented.

MAPOUT

is an output map with correlation coefficients at each grid point. There are two possible correlation coefficients:

1.	CC	[OUTPUT MAP]
2.	CC * ( 1.0 - sd(CC)/CC )	[OUTPUT SMAP]

This is the most important output: you can read it into PEAKMAX to extract peak positions which correspond to centers of your rotation axis. Or even better: use your favourite graphics program (e.g. O) and look for high peaks and/or long stretches of "density".

KEYWORDED INPUT

Available keywords are:

CHECK, END, INPUT, MINDEN, ORTHO, OUTPUT, POLAR, REPORT, SKIP, SLICE, SPHERE, STEP, XYZLIMIT

COMPULSORY KEYWORDS:

POLAR <omega> <phi> <kappa> [<omega-2> <phi-2> [<kappa-2>] ]

Polar angles of a selfrotation solution (definition as in POLARRFN).

Combining two selfrotation solutions:
If you have a twofold perpendicular to your rotation axis (e.g. D4 symmetry) you can give the polar angles as <omega-2> and <phi-2> (<kappa-2> defaults to 180.0). A corresponding sphere/disk will be built. The program stops if the two rotations aren't perpendicular. If they are perpendicular within an error of 5 degrees, the program calclulates a new 2-fold which now is exactly perpendicular, thus correcting possible rounding errors of e.g. POLARRFN.

ADDITIONAL KEYWORDS:

ORTHO <ncode>

Polar angles given on POLAR card are for orthogonalization code <ncode>.

ncode = orthogonalization code:

<ncode> =1: axes along a, c* x a, c* (Brookhaven standard, default)

<ncode> =2: axes along b, a* x b, a*

<ncode> =3: axes along c, b* x c, b*

<ncode> =4: axes along a+b, c* x (a+b), c*

<ncode> =5: axes along a*, c x a*, c ( Rollett )

<ncode> =6: axes along a, b*, a x b*

<ncode> =7: axes along a*, b, a* x b (TNT convention)

SPHERE <outer-radius> [<inner-radius>]

defines a spherical shape of your multimer.

Builds a sphere with radius <outer-radius>. You can omit a smaller inner sphere by giving <inner-radius>.

The sphere will be divided into <ifold> segments (where <ifold> is determined by <kappa>) and rotated so that its rotation axis is parallel with the selfrotation axis and its center is at (0 0 0). You can write this sphere out to logical XYZOUT.

To get a rough idea what your protein looks like: use the molecular weight Mr to get radius of assumed spherical protein:


                           1.23 * Mr * 0.75
               radius =  ( ---------------- ) ^ 0.333
                                  pi

default: <outer-radius>=25. <inner-radius>=0.

SLICE <outer-radius> <height> [<inner-radius>]

defines a different shape of your multimer.

Builds a disk with outer radius <outer-radius> and height <height>. You can omit a smaller inner circle by giving <inner-radius>.

The disk will be divided into <ifold> segments (where <ifold> is determined by <kappa>) and rotated so that its rotation axis is parallel with the selfrotation axis and its center is at (0 0 0). You can write this disk out to logical XYZOUT.

default: <outer-radius>=25. <height>=15. <inner-radius>=0.

CHECK [[NO]CORR] [[NO]PACK] [[N]AX1/[N]AX2/[N]AX3/[N]AX4]

which checks to perform:

[NO]CORR
[do not] calculate correlation coefficient at each search position

[NO]PACK
[do not] calculate amount of overlapping of all segments after applying all symmetry operators.

This didn't help a lot in my cases, but it's perhaps worth trying if your multimer covers a whole asymmetric unit.

[[N]AX1/[N]AX2/[N]AX3/[N]AX4]
after getting the correlation at each grid point, this correlation map is [not] searched for long stretches of high correlation by using points on the rotation axis (defined with keyword POLAR).

different weighting schemes are available:

AX1: CC = <CC>_axis(i)

AX2: CC = CC * <CC>_axis(i)

AX3: CC = CC * ( <CC>_axis(i) / CCmax )

AX4: CC = CC * ( <CC>_axis(j) / CCmax )

where:

CC	=	correlation at position i
CCmax	=	maximum correlation found
<CC>_axis(i)	=	average correlation for all points on rotation axis when centred at position i
<CC>_axis(j)	=	average correlation for all points i on rotation axis when centred at position j and weighted by 1/distance to position i

AX2 and AX3 don't make any difference in the result (but AX3 keeps the absolute values of the output correlation map at a reasonable height).

defaults: CORR NOPACK AX4

SKIP [AUTO <askip>]/[<iskip>]

Saves CPU time by using only a limit number of the points describing a sphere/slice.

Takes only every <iskip>th point of each segment in your sphere/disk to compute correlation coefficients. This is a good idea if your sphere/disk is rather big. It can save a lot of CPU. But take care that you keep at least ~500 points in each segment.

If keyword AUTO is present, the actual value of iskip is set so that approximately <askip> points per segment are used.

default: AUTO 500

STEP <istep>

Step along each cell axis (in grid units).

Unless you have calculated your map on a very fine grid, it does make things worse. And perhaps you'll miss the right solution !! It doesn't save a lot of CPU, since we have to interpolate the values at the end anyway.

default: <istep>=1

MINDEN <minden>

Correlation coefficients will only be calculated if the density for all segments in the sphere/disk is .gt. <minden>*sigma.

The default is also a very reasonable value.

default: <minden>=-999.

XYZLIMIT <xmin> <xmax> <ymin> <ymax> <zmin> <zmax>

Limits (in grid points) for search.

Unless you know already where to look for your multimer, I would always search the whole unit cell.

default: whole unit cell

OUTPUT [XYZ/GXYZ] [MAP/NOMAP] [SMAP]

MAP/SMAP = output MAPOUT

MAP = map with CC at each grid point

SMAP = map with CC*(1.-sd(CC)/CC) at each grid point (probably only useful with high symmetries: 4-fold,6-fold,D4,...)

NOMAP = no MAPOUT

XYZ/GXYZ = output XYZOUT

XYZ = orthogonal coordinates before interpolation

GXYZ = orthogonal coordinates after interpolation

default: MAP

INPUT XYZ

read in PDB file to define the shape of your molecules.

If you have a pretty good idea what your molecule looks like and how it is oriented (but not positioned) this could be quite helpful. But some restrictions:

different molecules/segments have to have different chain-ids

for each chain id there should be EXACTLY the same amount of atoms in exactly the same order

the multimer should be centred at the origin

REPORT <report> <top>

Not only reports the maximum correlation found so far, but also every correlation .gt. <report>.

At the end of the search the found correlations are sorted according to height and the <top> number is reported.

default: <report>=1. <top>=20

END

Terminates input.

EXAMPLES

A unix example script for performing a simple NCS search can be found in $CEXAM/unix/non-runnable/

getax-ncs-search

though it will need to be edited before use.

Other examples:

1. simple 2-fold

      getax mapin mlphare_6.0.map \
            mapout getax_sphere.map \
            <<end_ip >getax_sphere.log
      POLAR 51.7 90 180
      SPHERE 25.0
      END
      end_ip

      peakmax mapin getax_sphere.map \
              <<end_ip >getax_sphere.peakmax
      THRE RMS 4
      NUMP 100
      OUTP NONE
      end_ip

2. D4 symmetry

      getax mapin mlphare_6.0.map \
            mapout getax_slice.map \
            <<end_ip >getax_slice.log
      POLAR 48.7 116.7 90.0 90.0 28.4 180.0
      SKIP AUTO 1000
      SLICE 25.0 15.0 5.0
      REPORT 0.100
      CHECK NAX4
      END
      end_ip

      peakmax mapin getax_sphere.map \
              <<end_ip >getax_sphere.peakmax
      THRE RMS 4
      NUMP 100
      OUTP NONE
      end_ip

AUTHOR

Clemens Vonrhein

REFERENCES

C. Vonrhein and G. E. Schulz, Acta Cryst., D55, 225 - 229 (1999)
Locating proper non-crystallographic symmetry in low-resolution electron-density maps with the program GETAX.