Structural basis of transcription: alpha-amanitin-RNA polymerase II cocrystal at 2.8 A resolution

Abstract

The structure of RNA polymerase II in a complex with the inhibitor alpha-amanitin has been determined by x-ray crystallography. The structure of the complex indicates the likely basis of inhibition and gives unexpected insight into the transcription mechanism.

Figure 1

Stereo image of final α-amanitin structure. (A) σA-weighted F_obs − F_calc electron density at 2.8 Å resolution (red) contoured at 3 sigma calculated from the initial pol II placement before α-amanitin was included in the model. The final α-amanitin structure is shown (ball and stick model). (B) σA-weighted 2F_obs − F_calc electron density at 2.8 Å resolution (blue) contoured at 1.2 sigma, superimposed on the final α-amanitin structure (ball and stick model). Only the electron density around α-amanitin is shown. This figure was generated by using BOBSCRIPT and RASTER3D (–23).

Figure 2

Location of α-amanitin bound to pol II. (A) Cutaway view of a pol II-transcribing complex showing the location of α-amanitin binding (red dot) in relation to the nucleic acids and functional elements of the enzyme. Adapted from ref. . (B) Ribbons representation of the pol II structure (top view in refs. and 7). Eight zinc atoms are shown in light blue, the active site magnesium is magenta, the region of Rpb1 around α-amanitin is light green (funnel) and dark green (bridge helix), the region of Rpb2 near α-amanitin is dark blue, and α-amanitin is red. This figure was prepared by using RIBBONS (25).

Figure 3

Interaction of α-amanitin with pol II. (A) The chemical structure of α-amanitin, with residues of pol II that lie within 4 Å [determined by using CONTACT (26)] placed near the closest contact. The Cαs of α-amanitin are labeled with blue numbers. Hydrogen bonds are shown as dashed lines with the distances indicated. (B) Stereoview of the α-amanitin binding pocket. Ball and stick models of α-amanitin (red bonds) and of pol II residues within 4 Å (gray bonds) are shown. Rpb1 from A700 to A809 (funnel region) is light green. Rpb1 from A810 to A825 (bridge helix) is dark green. Rpb2 from B760 to B769 is blue. This figure was generated by using BOBSCRIPT and RASTER3D (–23).