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Figure 1.
Figure 1. Calculating an Experimental Electron Density Map of
Pol II by using Zn-MAD
(A) A representative anomalous
difference Fourier map (7.2 Å) based on a set of Zn peak
data and the model phases. Density contours are displayed at the
3.0σ (standard deviation) level in red. Three of the eight Zn
sites are shown as green spheres.
(B) A representative
dispersive difference Fourier map (7.2 Å) based on a set
of dispersive data and the model phases. The same map parameters
as in (A) were used in the calculation. The color scheme remains
the same as in (A).
(C) A representative crosscrystal
dispersive difference Fourier map (7.2 Å) used to assess
crystal isomorphism. The map was calculated by using the model
phases and amplitude differences between the remote data of
crystal-1 and the inflection data of crystal-2. The map
parameters and color scheme are the same as in (B).
(D–I)
(D) An experimental map based solely on Zn anomalous peak data
(SAS) and the known coordinates of the Zn sites. The map was
calculated to 7.2 Å and is shown in red at 1.0σ. An
existing 12 subunit Pol II model (Cα only) is shown in blue.
The green wire model is a lattice-packing neighbor. The
heterodimer of Rpb4/7 (indicated by “Rpb4/7”) protrudes to
the right from the bottom of the clamp domain (indicated by
“Clamp”). All of the maps in (D)–(I) were solvent
flattened as described in Experimental Procedures. Map slab
thickness, box sizes, and the color scheme are the same in
(D)–(I). (E) An experimental map calculated from a Zn
dispersive data set, treated as SIR. The resolution cutoff is
the same as in (D). (F) A Zn-MAD map (7.2 Å) resulted from
combining the anomalous and dispersive information. Lattice
contacts (indicated by “Contacts”) clarify in the full MAD
map, as indicated by the arrow. (G) The Zn-MAD map (7.2 Å)
produced from combining multiple Zn-MAD phasing sets. The Rpb8
β sheet (indicated by “Rpb8”) is labeled. (H) The Zn-MAD
map resulting from the manually edited mask (for solvent
flattening). The polymerase active site is indicated by the
arrow. (I) The final Zn-MAD map calculated to 4.1 Å. The
map was computed with smaller grid spacing than the
low-resolution maps described in other panels; thus, it appears
more crowded. Significantly more details of the polypeptide
chains become visible in this map. The arrow indicates the
density for fork loop-1 that was disordered in previous
uncomplexed Pol II structures.
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