Structures of a large prolate virus capsid in unexpanded and expanded states generate insights into the icosahedral virus assembly

Abstract

Many icosahedral viruses assemble proteinaceous precursors called proheads or procapsids. Proheads are metastable structures that undergo a profound structural transition known as expansion that transforms an immature unexpanded head into a mature genome-packaging head. Bacteriophage T4 is a model virus, well studied genetically and biochemically, but its structure determination has been challenging because of its large size and unusually prolate-shaped, ∼1,200-Å-long and ∼860-Å-wide capsid. Here, we report the cryogenic electron microscopy (cryo-EM) structures of T4 capsid in both of its major conformational states: unexpanded at a resolution of 5.1 Å and expanded at a resolution of 3.4 Å. These are among the largest structures deposited in Protein Data Bank to date and provide insights into virus assembly, head length determination, and shell expansion. First, the structures illustrate major domain movements and ∼70% additional gain in inner capsid volume, an essential transformation to contain the entire viral genome. Second, intricate intracapsomer interactions involving a unique insertion domain dramatically change, allowing the capsid subunits to rotate and twist while the capsomers remain fastened at quasi-threefold axes. Third, high-affinity binding sites emerge for a capsid decoration protein that clamps adjacent capsomers, imparting extraordinary structural stability. Fourth, subtle conformational changes at capsomers’ periphery modulate intercapsomer angles between capsomer planes that control capsid length. Finally, conformational changes were observed at the symmetry-mismatched portal vertex, which might be involved in triggering head expansion. These analyses illustrate how small changes in local capsid subunit interactions lead to profound shifts in viral capsid morphology, stability, and volume.

Assembly and Capsid Expansion Mechanism of Bacteriophage P22 Revealed by High-Resolution Cryo-EM Structures

“…The most dramatic change occurred in the coat N-arm domain, in which the N-terminal α-helix became the extending loop. A similar conformational change upon maturation also occurred in phages T4, T5, T7, SPP1, P23–45, and lambda [ 9 , 10 , 11 , 46 , 47 ] with different triangle numbers and capsid sizes ( Figure S12 ), suggesting that this structural change is conserved in the tailed phages, and is independent of the triangle number and capsid size. In addition, the tilts of the P-domain and E-loop were also conserved in these phages.…”

“…A similar conformational change in the A-domain also occurred in the coat proteins of phages SPP1, T5, and T7 [ 9 , 10 , 46 ] ( Figure S13 ). However, the local sixfold axes in both procapsids and capsids of phages T4, lambda, and HK97 [ 12 , 13 , 47 , 48 ] were closed ( Figure S14 ). Phages P22, SPP1, T7, T4, and lambda [ 3 , 9 , 10 , 49 , 50 ] encode scaffolding proteins for procapsid assembly.…”

Bacteriophage T4 Head: Structure, Assembly, and Genome Packaging

“…The structure and dimensions of the phage T4 prolate capsid shell are displayed in Figure 2 A. The head is elongated along its fivefold axis and has a length of 120 nm and a width of 86 nm [ 14 , 15 ]. The head encapsidates ~171 kbp linear double-stranded genomic DNA (~2–3% more than the unit-length genome).…”

“…Like the major capsid proteins of other tailed phages, gp23* subunits [ 15 , 18 ] have a polypeptide fold similar to that of the bacteriophage HK97 capsid protein [ 19 ] ( Figure 2 B). This fold is characterized by the wedge-shaped axial (A) domain located near the capsomer axis and the peripheral (P) domain, forming the capsomer’s periphery [ 19 ].…”

“…This fold is characterized by the wedge-shaped axial (A) domain located near the capsomer axis and the peripheral (P) domain, forming the capsomer’s periphery [ 19 ]. T4 gp23* has an additional 60-residue globular insertion (I) domain, which makes characteristic bumps on the capsid surface [ 15 , 18 , 20 , 21 ]. This I domain is connected to the rest of the structure via long linkers, which are analogous to the elongated E loop in the HK97 fold [ 19 ].…”