16. The map is colored according to the XY plane resolution.
Rotating the map automatically changes its color to reflect
the relative resolutions in the plane of view; it will also change
the relative Z-resolution curve in the second window. The user
can actively monitor these changes to determine the worst
resolution direction. An intuitive way to analyze these outputs
is to rotate the map to a point where the Z-resolution curve in
the second window reflects the lowest resolution. Once this is
accomplished, then the out-of-plane direction is the worst
resolved view; this direction is usually (but not necessarily)
also the “true” Z-direction relative to the electron beam for
this particular orientation (see Note 15). There could also be
multiple preferred orientations relative to the electron beam.
The out-of-plane view is not visible to the user, and therefore
the user cannot visually judge the detrimental effects on the
map in this orientation. Conversely, the view that is displayed
on the screen (the X/Y plane) should be the best resolved view.
Accordingly, the map should now be colored blue (the “best”
resolution in the X/Y plane). If the user now rotates the map
90
about the local X-axis, this would place the old XZ plane in
the field of view and the old Y direction out-of-plane. Now the
map should be colored red, because the “worst” resolution
(the Z-resolution relative to the electron beam) is now in the
field of view. Likewise, if the user rotates the map back, and
then 90
about the local Y-axis, this would place the old YZ
plane in the field of view, and the old X direction out of plane.
Again, the map should be colored red because the “worst”
resolution is still in the field of view. These procedures, and
how the Z-resolution and map coloring change with rotations,
are summarized in Fig. 4. Rotating the map around its different
axes, carefully monitoring the best and worst Z-resolution, and
inspecting the density will help to show how features are elongated in the Z-direction relative to the electron beam.
17. The radial points of the 3D FSC correspond to the Fourier
lattice points in the 3D transform, multiplied by the box size,
with units of inverse Angstroms (Å
À1 ). Therefore:
Spatial frequency in Å
À1 ¼ Radial point on 3DFSC
ð
Þ = Box size
ð
Þ :
The reason for the incorporation of the box size factor is to
make the numbers more user friendly. For example, if a map
has a voxel size of 0.79 Å, then the 3D FSC displays a value for
the voxel size as 1/0.79 ¼ 1.266, and it is easy to read off the
voxel values of the 3D FSC from the chimera plots. The
appropriate voxel size value is defined in the “Coordinates”
tab under “Volume Viewer” in Chimera. To convert the radial
lattice points of the 3D FSC into resolution (as displayed in
Local and Directional Resolution in Cryo-EM Maps
183
Rotating the map automatically changes its color to reflect
the relative resolutions in the plane of view; it will also change
the relative Z-resolution curve in the second window. The user
can actively monitor these changes to determine the worst
resolution direction. An intuitive way to analyze these outputs
is to rotate the map to a point where the Z-resolution curve in
the second window reflects the lowest resolution. Once this is
accomplished, then the out-of-plane direction is the worst
resolved view; this direction is usually (but not necessarily)
also the “true” Z-direction relative to the electron beam for
this particular orientation (see Note 15). There could also be
multiple preferred orientations relative to the electron beam.
The out-of-plane view is not visible to the user, and therefore
the user cannot visually judge the detrimental effects on the
map in this orientation. Conversely, the view that is displayed
on the screen (the X/Y plane) should be the best resolved view.
Accordingly, the map should now be colored blue (the “best”
resolution in the X/Y plane). If the user now rotates the map
90
about the local X-axis, this would place the old XZ plane in
the field of view and the old Y direction out-of-plane. Now the
map should be colored red, because the “worst” resolution
(the Z-resolution relative to the electron beam) is now in the
field of view. Likewise, if the user rotates the map back, and
then 90
about the local Y-axis, this would place the old YZ
plane in the field of view, and the old X direction out of plane.
Again, the map should be colored red because the “worst”
resolution is still in the field of view. These procedures, and
how the Z-resolution and map coloring change with rotations,
are summarized in Fig. 4. Rotating the map around its different
axes, carefully monitoring the best and worst Z-resolution, and
inspecting the density will help to show how features are elongated in the Z-direction relative to the electron beam.
17. The radial points of the 3D FSC correspond to the Fourier
lattice points in the 3D transform, multiplied by the box size,
with units of inverse Angstroms (Å
À1 ). Therefore:
Spatial frequency in Å
À1 ¼ Radial point on 3DFSC
ð
Þ = Box size
ð
Þ :
The reason for the incorporation of the box size factor is to
make the numbers more user friendly. For example, if a map
has a voxel size of 0.79 Å, then the 3D FSC displays a value for
the voxel size as 1/0.79 ¼ 1.266, and it is easy to read off the
voxel values of the 3D FSC from the chimera plots. The
appropriate voxel size value is defined in the “Coordinates”
tab under “Volume Viewer” in Chimera. To convert the radial
lattice points of the 3D FSC into resolution (as displayed in
Local and Directional Resolution in Cryo-EM Maps
183