Role of the Scaffolding Protein in P22 Procapsid Size Determination Suggested by T=4 and T=7 Procapsid Structures

Abstract

Assembly of bacteriophage P22 procapsids requires the participation of approximately 300 molecules of scaffolding protein in addition to the 420 coat protein subunits. In the absence of the scaffolding, the P22 coat protein can assemble both wild-type-size and smaller size closed capsids. Both sizes of procapsid assembled in the absence of the scaffolding protein have been studied by electron cryomicroscopy. These structural studies show that the larger capsids have T = 7 icosahedral lattices and appear the same as wild-type procapsids. The smaller capsids possess T = 4 icosahedral symmetry. The two procapsids consist of very similar penton and hexon clusters, except for an increased curvature present in the T = 4 hexon. In particular, the pronounced skewing of the hexons is conserved in both sizes of capsid. The T = 7 procapsid has a local non-icosahedral twofold axis in the center of the hexon and thus contains four unique quasi-equivalent coat protein conformations that are the same as those in the T = 4 procapsid. Models of how the scaffolding protein may direct these four coat subunit types into a T = 7 rather than a T = 4 procapsid are presented.

Structural basis for scaffolding-mediated assembly and maturation of a dsDNA virus

“…Although scaffolding proteins mediate the assembly of the procapsid shell, they are not required for maintaining the capsid shell, as evident in our empty procapsid structure (Fig. S4A) and the T ¼ 4 capsid structure (45), both of which show the skewed conformation for hexamers.…”

“…The lack of scaffolding proteins results in the failure to incorporate the portal and can lead to incomplete particles (29,32,40,45). Based on our density maps, we propose that the formation of a unique portal complex with the requisite scaffolding and coat proteins is likely the key for initiating proper procapsid assembly (Fig.…”

“…S4B) may explain how scaffolding proteins control the ratio between hexamers and pentamers during icosahedral shell assembly. In the absence of scaffolding proteins, the coat proteins sometimes polymerize into smaller T ¼ 4 shells (45). Both the T ¼ 4 and T ¼ 7 particles have 12 pentamers, though the T ¼ 4 particles have 30 skewed hexamers compared to 60 in T ¼ 7 particles.…”

Role of electrostatic interactions in the assembly of empty spherical viral capsids

“…T = h 2 + hk + k 2 and h and k are non-negative integers 11 . In view of the potential polymorphism of the viral protein assemblies which is also observed experimentally, especially in some viruses 6,12,13 , it is rather surprising that empty capsids of most simple viruses precisely assemble in capsids whose T -number is the same as in fully functional capsids that contain the viral RNA/DNA 2,3 . Even when several differently sized empty capsids do form 4,5,13 , these represent only a tiny subset of all of the imaginable capsid formations.…”

“…In view of the potential polymorphism of the viral protein assemblies which is also observed experimentally, especially in some viruses 6,12,13 , it is rather surprising that empty capsids of most simple viruses precisely assemble in capsids whose T -number is the same as in fully functional capsids that contain the viral RNA/DNA 2,3 . Even when several differently sized empty capsids do form 4,5,13 , these represent only a tiny subset of all of the imaginable capsid formations. Since the viral proteins have certain charge in the solution 15,16 , it is tempting to assume that the repulsive electrostatic interactions compete with some other attractive interactions between the viral proteins in such a way that the total free energy of the capsid is minimal exactly for the capsid of the observed radius.…”