Data”. From the same menu one might also use the “Segment
Map” option [11]. In addition, if a preliminary atomic model is
available for a body, then the “molmap” command in UCSF
Chimera can be used to convert the fitted PDB into a density
map (see https://www.cgl.ucsf.edu/chimera/docs/
UsersGuide/midas/molmap.html); and the volume operation (“vop”) option “resample onGrid” (see https://www.
cgl.ucsf.edu/chimera/docs/UsersGuide/
framecommand.html) can be used to generate a map on the
same Cartesian grid as the consensus map. This map can then
be saved from the “Volume Viewer” window, and input into
the “Mask creation” job-type of RELION. Whatever the
method chosen, make sure the final masks do not have highresolution features and only have values in the range [0,1] by
applying low-pass filtering and soft-edge addition in the “Mask
creation” job-type.
5. If the definition of a body smaller than 150 kDa is necessary, it
is possible to fix the relative orientation of this body by defining
the widths of its rotational and translational priors to zero.
6. Within a typical RELION project, the optimiser.star file is
directly visible through the “Browse” button, but this is not
the case in this simplified example. Therefore, just type its
location in the GUI text entry.
7. The value of the offset range specifies the maximum translational offsets searched in every iteration. As the translational
searches are centred at the optimal translation from the previous iteration, individual particles can shift more than this search
range during multiple iterations. Therefore, this value can be
smaller than the total search allowed by the prior on the translations as specified in step 5, Subheading 3.1.
8. The ten maps that are generated for each principal motion in
the data will have relative orientations of their bodies that
correspond to the median orientations of ten equi-populated
subsets of particles along the principal motion.
9. By setting “Select particles based on eigenvalues?
Yes” on the “Analyse” tab, one can generate STAR files
with subsets of the particles that have eigenvalues above or
below the specified minimum or maximum values for any of
the principal components, as specified on the GUI. This calculation is run as a continuation of a previously performed multibody job, by pressing the “Continue!” button. Multiple
selections can be performed in a single job directory. The
output STAR files with the selected subsets can then be used
for further 3D classification or refinement.
Multi-body Refinement in RELION
155
Map” option [11]. In addition, if a preliminary atomic model is
available for a body, then the “molmap” command in UCSF
Chimera can be used to convert the fitted PDB into a density
map (see https://www.cgl.ucsf.edu/chimera/docs/
UsersGuide/midas/molmap.html); and the volume operation (“vop”) option “resample onGrid” (see https://www.
cgl.ucsf.edu/chimera/docs/UsersGuide/
framecommand.html) can be used to generate a map on the
same Cartesian grid as the consensus map. This map can then
be saved from the “Volume Viewer” window, and input into
the “Mask creation” job-type of RELION. Whatever the
method chosen, make sure the final masks do not have highresolution features and only have values in the range [0,1] by
applying low-pass filtering and soft-edge addition in the “Mask
creation” job-type.
5. If the definition of a body smaller than 150 kDa is necessary, it
is possible to fix the relative orientation of this body by defining
the widths of its rotational and translational priors to zero.
6. Within a typical RELION project, the optimiser.star file is
directly visible through the “Browse” button, but this is not
the case in this simplified example. Therefore, just type its
location in the GUI text entry.
7. The value of the offset range specifies the maximum translational offsets searched in every iteration. As the translational
searches are centred at the optimal translation from the previous iteration, individual particles can shift more than this search
range during multiple iterations. Therefore, this value can be
smaller than the total search allowed by the prior on the translations as specified in step 5, Subheading 3.1.
8. The ten maps that are generated for each principal motion in
the data will have relative orientations of their bodies that
correspond to the median orientations of ten equi-populated
subsets of particles along the principal motion.
9. By setting “Select particles based on eigenvalues?
Yes” on the “Analyse” tab, one can generate STAR files
with subsets of the particles that have eigenvalues above or
below the specified minimum or maximum values for any of
the principal components, as specified on the GUI. This calculation is run as a continuation of a previously performed multibody job, by pressing the “Continue!” button. Multiple
selections can be performed in a single job directory. The
output STAR files with the selected subsets can then be used
for further 3D classification or refinement.
Multi-body Refinement in RELION
155