Structure of the C-terminally truncated P22 portal. (A) The map was rotated by 45° about the horizontal axis sequentially from the left to the right. (B) Cutaway view. The three domains are outlined by dashed ellipses. (C) Fitting of the X-ray structure of phi29 portal (the ribbon diagram) into the cryoEM map of the P22 portal (semitransparent isosurface). Each subunit of the phi29 portal is in a unique color. Only the front half is shown for clarity.

Structure of the P22 tail machine. (A) Cutaway view of the P22 tail machine (blue isosurface). The cryoEM map of C-terminally truncated portal (red isosurface) is superimposed. The X-ray structures of the receptor-binding domains and head-binding domains of the homotrimeric tailspike (ribbon diagrams with each polypeptide in red, green and yellow) are docked into the map. The density for gp4 and gp10 is indicated by a dashed curve, and the putative boundary between them by a dotted line. (B) Cross-section of the tail machine map at the position indicated by a dotted arrow in panel A. The atomic model of the receptor-binding domains (ribbon diagrams with each polypeptide in red, green and yellow) is superimposed with one of six tailspike trimers. A contour map (density value from 4.35σ to maximum with an interval of 1.74σ) is superimposed to show the trimeric conformation of the tailspike. The tiny feature (asterisk) in the middle of each tailspike represents lower density. (C) A contour (density value from 4.35σ to maximum with an interval of 0.58σ) of a cross-section in the tail machine map corresponding to the portal. The position of the cross-section is indicated by a dashed arrow in panel A. (D) The bottom opening of the tail tube viewed from the gp26 needle. The density that connects gp26 and gp10 is indicated by red arrows.

The assembly cascade of the P22 tail machine, starting from the phage capsid with the portal incorporated in a unique five-fold vertex (A), followed by sequential attachment of gp4 (B), gp10 (C), gp26 (D) and tailspikes (E). The tail machine is putatively superimposed with a part of a cryoEM map (gray isosurface) of the wild-type P22 (L Tang, JE Johnson and SR Casjens, unpublished data). The front half of the structures in (A–E) was computationally removed to show the internal features. A front view of the putative P22 structure is shown in (F).

Electron micrographs of the frozen-hydrated P22 tail machine. (A) Representative view of the electron micrograph. Inset: a class average. (B) CCD images over the same field taken at tilt angles of 0, 20 and 40°. (C) Tilt series by 0, 20, 30, 40 and 60° (from left to right). Bars: 200 Å.

CryoEM reconstruction of the P22 tail machine. The map was rotated by 45° along the horizontal axis stepwise from left to right. The map was contoured at 3.7σ above background and the resultant volume corresponds to 100% of the mass.

Secondary structure prediction of gp26 by PHD (http://www.predictprotein.org/) (Rost et al, 2004). The two putative heptad repeat regions are indicated by red and green background, and the characteristic first and fourth hydrophobic residues in each repeat unit are indicated beneath the sequence by letters a and d, respectively. The secondary structure is labeled by blue cylinders for α helix and light blue arrows for β strand.

Fourier shell correlation plots of reconstructions of (A) the truncated P22 portal and (B) the P22 tail machine. In (B), the Fourier shell correlation drops to 0.5 at 1/23 Å⁻¹, 0.4 at 1/22 Å⁻¹, and 0.3 at 1/16 Å⁻¹.