Subunit tomato bushy stunt virus

Figure 16.8 Architecture of the tomato bushy stunt virus particle. The polypeptide chain of each subunit folds into three domains (R, S, P) with a 35-residue connecting arm (a) between R and S and a hinge (h) between S and P.

The size of this viral particle is of course larger than that of a virus with only 60 subunits. The diameter of tomato bushy stunt virus is 330 A compared with 180 A for satellite tobacco necrosis virus. The increase in volume of the capsid means that a roughly four times larger RNA molecule can be accommodated. 

The N-terminal part of the tomato bushy stunt virus polypeptide chain (the R-segment in Figure 16.8) is disordered in all the subunits. As in the core of many other single-strand RNA viruses this region of the polypeptide chain... 

Figure 16.9 Contacts between P domains in tomato bushy stunt virus. Sa, Sb, and Sc are the shell domains of subunits A, B, and C, respectively. Pa, Pb, and Pc are the protruding domains of subunits A, B, and C, respectively.

Figure 16.10 The arms of all 60 C subunits in tomato bushy stunt virus form an internal framework, (a) Configuration of interdigitated arms from the three C subunits, viewed down a threefold axis. The p strands are shown as arrows, (b) Cutaway view of the virus particle, emphasizing the framework function of the C-subunit arms. These arms are shown as chains of small balls, one per residue. The region where three arms meet and interdigitate is shown schematically in (a). The main part of each subunit is represented by large balls. Only about one hemisphere of these is drawn, but all the C-subunit arms are included.

The asymmetric unit contains one copy each of the subunits VPl, VP2, VP3, and VP4. VP4 is buried inside the shell and does not reach the surface. The arrangement of VPl, VP2, and VP3 on the surface of the capsid is shown in Figure 16.12a. These three different polypeptide chains build up the virus shell in a way that is analogous to that of the three different conformations A, C, and B of the same polypeptide chain in tomato bushy stunt virus. The viral coat assembles from 12 compact aggregates, or pen tamers, which contain five of each of the coat proteins. The contours of the outward-facing surfaces of the subunits give to each pentamer the shape of a molecular mountain the VPl subunits, which correspond to the A subunits in T = 3 plant viruses, cluster at the peak of the mountain VP2 and VP3 alternate around the foot and VP4 provides the foundation. The amino termini of the five VP3 subunits of the pentamer intertwine around the fivefold axis in the interior of the virion to form a p stmcture that stabilizes the pentamer and in addition interacts with VP4. 

One of the most striking results that has emerged from the high-resolution crystallographic studies of these icosahedral viruses is that their coat proteins have the same basic core structure, that of a jelly roll barrel, which was discussed in Chapter 5. This is true of plant, insect, and mammalian viruses. In the case of the picornaviruses, VPl, VP2, and VP3 all have the same jelly roll structure as the subunits of satellite tobacco necrosis virus, tomato bushy stunt virus, and the other T = 3 plant viruses. Not every spherical virus has subunit structures of the jelly roll type. As we will see, the subunits of the RNA bacteriophage, MS2, and those of alphavirus cores have quite different structures, although they do form regular icosahedral shells. 

Figure 16.13 The known subunit structures of plant. Insect, and animal viruses are of the jelly roll antiparallel p barrel type, described in Chapter 5. This fold, which is schematically illustrated in two different ways, (a) and (b), forms the core of the S domain of the subunit of tomato bushy stunt virus (c). [(b), (c) Adapted from A.J. Olson et al., /. Mol. Biol. 171 61-93, 1983.1...

Tomato bushy stunt virus is a T = 3 plant virus with 180 chemically identical subunits. Each polypeptide chain is divided into several domains. The subunits preserve quasi-equivalent packing in most contact regions by conformational differences of the protein chains, especially a large change in... 

One of the most intriguing recent examples of disordered structure is in tomato bushy stunt virus (Harrison et ah, 1978), where at least 33 N-terminal residues from subunit types A and B, and probably an additional 50 or 60 N-terminal residues from all three subunit types (as judged from the molecular weight), project into the central cavity of the virus particle and are completely invisible in the electron density map, as is the RNA inside. Neutron scattering (Chauvin et ah, 1978) shows an inner shell of protein separated from the main coat by a 30-A shell containing mainly RNA. The most likely presumption is that the N-terminal arms interact with the RNA, probably in a quite definite local conformation, but that they are flexibly hinged and can take up many different orientations relative to the 180 subunits forming the outer shell of the virus particle. The disorder of the arms is a necessary condition for their specific interaction with the RNA, which cannot pack with the icosahedral symmetry of the protein coat subunits. 

Fig. 69. The two different positions of the hinge between domains 2 and 3 of tomato bushy stunt virus protein. Each domain is represented by a cylinder, with domain 2 as the reference and domain 3 shown in the relative positions it takes in type B subunits and in type C subunits.

Fig. 70. Domains 2 (cylinders) and N-terminal tails of B- and C-type subunits around the quasi-6-fold axis in tomato bushy stunt virus. The association of the three C-subunit tails around the quasi-6-fold forms domain 1 (see Fig. 84).

The remaining three antiparallel /3 structures form a miscellaneous category (see Fig. 84). Lactate dehydrogenase d2 and gene 5 protein each has several two-stranded antiparallel j8 ribbons, but they do not coalesce into any readily described overall pattern. The N-terminal domain of tomato bushy stunt virus protein has a unique /3 structure in which equivalent pieces of chain from three different subunits wrap around a 3-fold axis to form what has been called a /3 annulus (Harrison et ah, 1978). Each of the three chains contributes a short strand segment to each of three three-stranded, interlocking /3 sheets. This domain provides one of the subunit contacts that hold the virus... 

The structure of the capsid (protein shell) for an icosahedral virus such as tomato bushy stunt virus. Pentons (P) are located at the 12 vertices of the icosahedron. Hexons (H), of which there are 20, form the edges and faces of the icosahedron. Each penton is composed of five protein subunits and each hexon is composed of six protein subunits. In all, the structure contains 180 protein subunits. 

In tomato bushy stunt virus, 180 identical protein subunits (Mr = 41,000) form a shell which surrounds a molecule of RNA containing 4,800 nucleotides. For geometrical reasons no more than 60 identical subunits can be positioned in a spherical shell in a precisely symmetrical way. This limits the volume, so, in order to accommodate a greater amount of RNA, a larger shell is needed. This can be achieved by relaxing the symmetry the subunits are divided into three sets of 60, each set packing with strict symmetry. However, the relationship between each set is different so that the overall packing is only quasi-equivalent. 

A single mechanism will not describe even the "controlled aggregation of proteins in nature. A single viral coat protein, for example, may form several different contacts with itself and other proteins, depending on its final position within the shell structure. Indeed, the original postulate of "quasi-equivalent binding at lattice points in virus capsules was modified to "non-equivalency" when the first structure was solved at atomic dimensions. For example, the coat of Tomato bushy stunt virus consists of 180 identical subunits of a 43 kD protein which self-interact at lattice points in at least three distinct ways k... 

In collaboration with the Virology Laboratory of the Institut de Biologie Moleculaire et Cellulaire in Strasbourg, the Laboratory of Molecular Acoustics has studied the ultrasonic absorption of two small icosahedral viruses, bromegrass mosaic virus (BMV) and tomato bushy stunt virus (TBSV). Measurements carried out between 0.6 and 40 MHz showed that ultrasonic waves in the MHz range are absorbed much more by capsids than by dissociated protein. Examples of ultrasonic spectra of solutions of BMV protein capsids and of subunit-dimers are shown in Fig. 1 (at 5 mg/ml in 10 mM Na cacodylate, 1.00 M NaCl, at 23°C capsids , pH=4.80 dimers o, pH = 6.6o). Spectra obtained for BMV (A, pH = 5 00, at the same particle concentration as for the BMV capsids, otherwise similar conditions) and for the pure solvent ( — ) are also shown. 

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