Condensed genome structure

Abstract

Large, tailed dsDNA-containing bacteriophage genomes are packaged to a conserved and high density (∼500 mg/ml), generally in ∼2.5-nm, duplex-to-duplex, spaced, organized DNA shells within icosahedral capsids. Phages with these condensate properties, however, differ markedly in their inner capsid structures: (1) those with a naked condensed DNA, (2) those with many dispersed unstructured proteins embedded within the DNA, (3) those with a small number of localized proteins, and (4) those with a reduced or DNA-free internal protein structure of substantial volume. The DNA is translocated and condensed by a high-force ATPase motor into a procapsid already containing the proteins that are to be ejected together with the DNA into the infected host. The condensed genome structure of a single-phage type is unlikely to be precisely determined and can change without loss of function to fit an altered capsid size or internal structure. Although no such single-phage condensed genome structure is known exactly, it is known that a single general structure is unlikely to apply to all such phages.

Fig. 21.1

Examples of different internal capsid arrangements found in tailed dsDNA bacteriophages. Tails on capsids are not shown. Portal complex depicted by purple; 2.5-nm spaced DNA depicted by shades of blue. (a) lambda, HK97, and Phi29 have only a “naked” condensed DNA within their capsids; (b) the T4 capsid contains more than a thousand dispersed internal proteins (yellow) embedded within its DNA, whose copy numbers vary relative to capsid dimensions, as well as several localized proteins whose copy numbers remain unchanged in different-sized capsids (red), such as icosahedral capsids (T4 i) or giant capsids (T4 g); (c) the T7 capsid has an inner core composed of three different proteins (yellow); and (d) the PhiKZ capsid contains an extremely large inner protein body (purple)

Fig. 21.2

Different models of packaged DNA structure. (a) The spool model containing longitudinally packaged DNA segments (Earnshaw et al. 1978); (b) spool or concentric shell models (Earnshaw and Harrison 1977; Cerritelli et al. 1997); (c) spiral-fold model (Black et al. 1985); (d) liquid crystal and hexagonal packing models (Lepault et al. 1987; Leforestier and Livolant 2010); (e) icosahedral-bend model which is the same as in (a) but sharp bends have been incorporated to follow the angular contours of the icosahedrons (Mullaney and Black 1998); (f) a model where the DNA forms 12 icosahedrally arranged pear-shaped rings was proposed for the large Bacillus phage G (Sun and Serwer 1997). This model is based on that proposed for lambda DNA (Witkiewicz and Schweiger 1985); (g) model for completion of genome packaging into shells in Phi29 based on a Monte Carlo simulation (Comolli et al. 2008); and (h) model for genome packaging with extensively knotted DNA that may arise following packaging (Marenduzzo et al. 2009). Nb. Capsid shell is not indicated in (b, f, g, and h)

Fig. 21.3

(a) Overview of the packaging of proteins by the T4 capsid targeting sequence (CTS) system: (i) capsid-size prohead core, minus the outer shell and DNA, is seen to assemble in vivo; (ii) completion of the capsid shell triggers cleavage of prohead proteins; (iii) small fragments of processed prohead proteins (including scaffold protein, CTS sequences, and protease) exit the capsid presumably via small holes in the shell prior to maturation–expansion; (iv) phage DNA is packaged into the capsid by the terminase; (v) mature T4 head. (b) Examples of proteins packaged into the mature T4 phage capsid: (i) structure of restriction endonuclease inhibitor IPI* (9 kDa) (the processed form of IPI) found in wild-type T4; examples of proteins packaged into the phage capsid using the phage-derived expression, packaging, and processing (PEPP); (ii) SN nuclease (15 kDa) (Chen et al. 2000); (iii) GFP (27 kDa) (3ADF) (Ebisawa et al. 2010); (iv) β-galactosidase (540-kDa tetramer) (1DP0) (Juers et al. 2000) (Nb. Structures not to scale). (c) Genomic DNA of recombinant T4 phage digested by SN nuclease within the capsid. Lane 1, lambda Hin dIII standard; lane 2, control DNA from recombinant T4[CTS ▼IPIII▼SN], not treated with Ca²⁺; lanes 3–7, DNA from recombinant T4[CTS▼IPIII▼SN] incubated with Ca²⁺ for 1, 10, 30, 120 min, and 16 h, respectively (▼refers to a T4 protease processing site). Image reproduced from Mullaney and Black (1998)

Fig. 21.4

The inner body found in the large myovirus, PhiKZ, and its relative 201phi2-1. (a) Electron micrograph of disrupted heads of the PhiKZ mutant ts759. Image reproduced from Krylov et al. (1984); (b) electron micrograph of partially disrupted PhiKZ particles. Arrow indicates the inner body. Image reproduced from Matsko et al. (2001); (c) electron micrograph of a PhiKZ particle disrupted by freeze–thawing. Arrow indicates inner body released from disrupted head. Image reproduced from Krylov et al. (1984); (d) cryo-electron micrograph of 201phi2-1. The large white arrow highlights bubbles which formed in the head apparently around the protein body. Image reproduced from Thomas et al. (2008)