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Nma_alignment_v3
This is a method for iterative elastic 3D-to-2D alignment that computes elastic and rigid-body parameters of the alignment of single-particle images with a reference structure through reference-structure deformations along normal modes followed by its rigid-body alignment with images. The elastic parameters are deformation amplitudes along the normal modes and the rigid-body parameters are three Euler rotations and two in-plane shifts.
The reference structure can be of atomic resolution (e.g., solved by X-ray crystallography) or pseudo-atomic resolution (e.g., obtained by electron microscopy (EM) and converted into a network of Gaussian functions) and should be given in the PDB format.
The conversion from an EM volume into a network of Gaussian functions can be done either on the webserver 3DEM Loupe or with xmipp_volume_to_pseudoatoms ( 3DEM Loupe actually runs xmipp_volume_to_pseudoatoms, which provides the pseudo-atomic structure in the PDB format required by xmipp_nma_alignment and xmipp_mpi_nma_alignment).
The normal modes can be computed using the webserver ElNemo if the reference structure has an atomic resolution or using the webserver 3DEM Loupe if the reference structure is an EM volume.
By default, the rigid-body alignment is done using a combination of a “discrete” alignment (maximizing the cross-correlation between the experimental image and a discrete library of reference projections in wavelet space for a robustness to noise) and a “continuous” alignment (gradient-based optimization of the similarity between the experimental image and central slices of the reference structure in Fourier space, which is used for a fast refinement of an initial “discrete” alignment). The wavelet-space projection matching can optionally be replaced by a real-space projection matching.
Align images with an atomic or pseudo-atomic (from EM volume) structure by computing deformation amplitudes along normal modes, three Euler angles, and two in-plane shifts
Parameters
--mode <modeoverwrite> where <mode> can be:
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- $
--label <image_labelimage> $-o, --output <output_file> $: Reference atomic or pseudo-atomic structure in PDB format $`--odir <outputDir`.> $: Resume processing
Generation of the deformed reference volumes $: File with a list of mode filenames $`--sampling_rate <Ts`1> $`--filterVol <cutoff`15.> $: Center the PDB structure $--fixed_Gaussian <std-1>
Combined elastic and rigid-body alignment $--trustradius_scale <s1> $--mask <m> --discrAngStep <ang10>
This is an example of running the method on a series of images (listed in selimg.sel) using a reference structure in PDB format (reference.pdb) that is centered (--centerPDB), modes listed in modelist.txt, pixel size of 3 Angstroms (--sampling_rate 3), angular step of 10° (--discrAngStep 10) for computing a discrete library of reference projections, and writing the results in output.txt.
mpiexec.hydra -machinefile $XMIPP_MACHINEFILE -np 8 ~/xmipp/bin/xmipp_mpi_nma_alignment -i selimg.sel --pdb reference.pdb --modes modelist.txt --sampling_rate 3 --discrAngStep 10 -o output.txt --centerPDB
In this example, we used MPI-parallelized version of the method to dispatch the jobs on 8 cores (-np 8) listed in the text file specified by the environment variable $XMIPP_MACHINEFILE (-machinefile $XMIPP_MACHINEFILE). Note that depending on your MPI version (e.g., if you use mpirun or mpiexec), you will have to adapt the first part of the command (mpiexec.hydra -machinefile $XMIPP_MACHINEFILE -np 8). Also, you have usually to provide the full path to the executable xmipp_mpi_nma_alignment (~/xmipp/bin/xmipp_mpi_nma_alignment). Here is how the required files should look like:
modelist.txt - List of text files containing the normal modes to be used (3-column text files). Here, we give an example of the file containing 2 modes:
# XMIPP_STAR_1 * # data_ loop_ _NMAModefile ../../modes_columns/mod0011 1 ../../modes_columns/mod0012 1
selimg.sel - List of images to analyze. Here, we give an example of the beginning of a file:
# XMIPP_STAR_1 * # data_noname loop_ _image _enabled images_simulated/img_pop1_000001.xmp 1 images_simulated/img_pop1_000002.xmp 1 images_simulated/img_pop1_000003.xmp 1 images_simulated/img_pop1_000004.xmp 1 images_simulated/img_pop1_000005.xmp 1 … …
Here is how the output file should look like:
output.txt – Columns before quotation marks: image name (1st), enabled (2nd), Euler angles (3rd-5th), in-plane shifts (6th-7th). In quotation marks: as many deformation amplitudes as the number of modes. Last column is the measure of the alignment quality. Here, we give an example of the beginning of a file from an experiment with 2 modes:
# XMIPP_STAR_1 * # data_noname loop_ _image _enabled _angleRot _angleTilt _anglePsi _shiftX _shiftY _nmaDisplacements _cost images_simulated/img_pop1_000004.xmp 1 -16.630944 -50.722823 19.072670 -2.027733 -2.121395 ' -246.744000 54.542600 ' 0.096851 images_simulated/img_pop1_000003.xmp 1 -5.324611 -13.784008 16.925189 2.256479 -1.727479 ' 252.391000 53.157900 ' 0.156012 images_simulated/img_pop1_000005.xmp 1 -1.604509 31.749473 -0.143788 -4.124537 -4.483128 ' -305.762000 293.316000 ' 0.025739
In case of a pseudo-atomic reference structure, the command to launch xmipp_mpi_nma_alignment differs from the one for an atomic-resolution structure only in using the flag --fixed_Gaussian (that was not used with the atomic reference structure) and in not using the flag --centerPDB. Here is an example of the command using the reference structure with Gaussian functions whose standard deviation is 2 (--fixed_Gaussian 2). The standard deviation of the Gaussian functions is given by the user at the time the volume is converted into Gaussian-type pseudo-atoms.
mpiexec.hydra -machinefile $XMIPP_MACHINEFILE -np 8 ~/xmipp/bin/xmipp_mpi_nma_alignment -i selimg.sel --pdb reference.pdb --modes modelist.txt --sampling_rate 3 --discrAngStep 10 -o output.txt --fixed_Gaussian 2
The easiest is to compute the pseudo-atomic structure with the 3DEM Loupe webserver that runs the code of xmipp_volume_to_pseudoatoms, and at the same time allows computing the normal modes for the structure.
Here is how the pseudo-atomic PDB-format file should look like:
REMARK xmipp_convert_vol2pseudo REMARK fixedGaussian 2.000000 REMARK intensityColumn Bfactor ATOM 1 DENS DENS 1 36.000 33.000 -58.350 1 0.20 DENS ATOM 2 DENS DENS 2 -91.350 22.500 49.650 1 0.26 DENS ATOM 3 DENS DENS 3 -58.050 0.000 -81.000 1 0.19 DENS ATOM 4 DENS DENS 4 -25.500 94.500 -7.500 1 0.17 DENS ATOM 5 DENS DENS 5 13.650 34.500 10.500 1 0.29 DENS ATOM 6 DENS DENS 6 82.650 34.650 -28.350 1 0.28 DENS …. ….
The remark “fixedGaussian 2.000000” means that the Gaussian functions have the standard deviation of 2. The remark “intensityColumn Bfactor” means that the amplitude of each Gaussian functions are written in the Bfactor column of the PDB-format file, which is mandatory for the elastic 3D-to-2D alignment method.
The radius of the trust region is a typical distance between successive points at which the objective function is evaluated. The radius is reduced automatically as soon as the objective function stops decreasing. The method is initialized by specifying the initial deformation amplitudes (they are set to zero which corresponds to the non-deformed reference structure) and the initial trust region radius (set to 250) and the final trust region radius (set to 0.5). The initial trust region radius can be scaled by specifying the scaling factor on the command line using --trustradius_scale(a default value of the scaling factor is 1).
Here is an example of using the scaling factor of 1.5 for an atomic structure:
mpiexec.hydra -machinefile $XMIPP_MACHINEFILE -np 8 ~/xmipp/bin/xmipp_mpi_nma_alignment -i selimg.sel --pdb reference.pdb --modes modelist.txt --sampling_rate 3 --discrAngStep 10 -o output.txt --trustradius_scale1.5 --centerPDB
and, for a pseudo-atomic structure:
mpiexec.hydra -machinefile $XMIPP_MACHINEFILE -np 8 ~/xmipp/bin/xmipp_mpi_nma_alignment -i selimg.sel --pdb reference.pdb --modes modelist.txt --sampling_rate 3 --discrAngStep 10 -o output.txt --fixed_Gaussian 2 --trustradius_scale1.5
xmipp_nma_alignment -i images.sel --pdb 2tbv.pdb --modes modelist.xmd --trustradius_scale 1.2 --sampling_rate 3.2 -o output.xmd --resume