1

Lecture 3. Physical Structures

Terms•Capsid•Envelopes•Nucleocapsid

•Helical•Icosahedral

Great website: http://viperdb.scripps.edu/

2

Size, mass, dimensions of viruses Particle- An aggregate of many molecules often of

different nature.

•Weights range from 3-800 X 106 Daltons (Da). •Linear dimensions generally given in nm (10-9 meters). Typical viruses range from ≈ 25 - >100 nm diameter. •Comparison: Bacterial cells: ≈1500 nm dia; Eukaryotic epithelial cells about ≈20,000 nM dia. •Assuming viruses and bacteria are nearly spherical (not always the case), a bacteria has a volume about 30,000 times greater than a virus while a epithelial cell is about 60 million times larger. could stuff about 3 x 1014 virus particles in one drop of water.

3

Size, mass, & dimensions

• S-value- A value derived from the sedimentation rate of particles and molecules in the ultracentrifuge.

• Numbers are reproducible under specific conditions and reflect the volume and shape of a particular substance but are not directly proportional to mass.

• The basic unit is the Svedberg (S) which is 10-

13 sec. • This value can be used to estimate molecular

weights in conjunction with other values.

4

Structural Principles and terms

• Helical-Rod or threadlike appearance• Isometric-spherical appearance• Irregular-without clear symmetry

5

Virus makeup• Protein subunit: individual folded protein molecules. • Structural subunit (synonyms; protomer, asymmetric unit): Unit from which capsid of

nucleocapsid are built; may comprise one protein subunit or multiple different subunits.• Morphological unit (syn: capsomere): surface structure (knobs, projections, clusters, etc.)

seen by electron microscopy. This term is generally restricted to descriptions of viruses from electron micrographs.

• Capsid (syn: coat): regular, shell-like structure composed of aggregated protein subunits which surrounds the viral nucleic acid.

• Nucleocapsid (syn: core): viral nucleic acid enclosed by a capsid protein coat.• Envelope (syn: viral membrane): lipid bylayer containing viral glycoproteins. The

phospholipids in the bylayer are derived from the cell that the virus arose from. Not all viruses have envelopes some consist of only the nucleocapsid.

• Virion: physical virus particle. Nucleocapsid alone for some viruses (picornaviruses) or including outer envelope structure for others (retroviruses).

6

The Viral Capsid• Capsid- Protein coat that encapsidates the virus.• Nucleocapsid-Capsid with genome inside (plus anything else

that may be inside like enzymes and other viral proteins for some viruses).

Capsid functions1. Protect genome from outside environment (May include

damaging UV-light, shearing forces, nucleases either leaked or secreted by cells).

2. Virus-attachment protein- interacts with cellular receptor to initiate infection. Since viruses are made of many different repeated subunits there is redundancy; Many receptor sites so damage to a few doesn’t prevent infection.

3. Delivery of genome in infectious form. May simply “dump” genome into cytoplasm (most +ssRNA viruses) or serve as the core for replication (retroviruses and rotaviruses).

7

How do particles form?• Information is encoded in the components themselves (nucleic acid

+ proteins). • Some proteins can form capsid shells in the absence of the genome;

others form around the genome. • Fraenkel-Conrat and Williams (1955): TMV nucleocapsids form

spontaneously from individual protein subunits (coat protein) and the genome.

Particles represent a free energy minimum state which leads to stability.

• Assembly is driven by hydrophobic and hydrophilic (rarely covalent) interactions including– Protein-protien, – Protein-nucleic acid– Protein-lipid

• Important: since particles must disassemble at some point during infections, covalent bonds would make this more difficult.

8

• Why not make the capsid from a single large protein rather than assemble it from many proteins?

• Not enough genomic information:• MW of 1 codon (3 nucleotides) is about 1000 Da. 1

amino acid is about 150 Da, a genome can only produce proteins that are 15% of its molecular weight.

• A picornavirus is about 10,000 bases----can produce a protein about 500,000 Dal.

• Outer shell of picornaviruses is make up of 60 copies each of 4 different proteins, approx MW=2 million Da.

• The genome does not have enough information to encode a single protein that could encapsidate it.

9

Virus Shapes

• Most viruses have evolved to form one of 2 different shapes;

• Helical and Icosohedral. • Some irregular viruses do exits and many of these

have underlying helical or icosohedral symmetry. • Note-viruses form regular shapes but use irregular

proteins to do so. This creates a problem that must be solved for assembly to occur. For example, it would be easy to imagine how a virus might form an icosahedron if perfectly triangular proteins were used. But the proteins are irregular shaped and still must form a sealed (to protect genome) icosahedron.

10

Helical nucleocapsidsTopology follows the biophysical geometry of the nucleic acid genome.

Enveloped Helicalnucleocapsid Schematic

representationof Tobacco Mosaic Virus.

EM of Influenza Cfilamentous particle

11

Helical viruses• Many biological components have adopted a helical structure

(DNA, a helix of proteins). • Energetically favorable and can allow flexibility (bend but don’t

break (shear)). • The simplest way to arrange irregular identical proteins would be

around a central axis to form a disk. Disks could then be stacked with the genome in the middle to form a cylinder.

• Helical viruses form a closely related spring like helix instead. The best studied TMV but many animal viruses and phage use this general arrangement. – Note-all animal viruses that are helical are enveloped, unlike many of the

phage and plant viruses. • Most helixes are formed by a single major protein arranged with

a constant relationship to each other (amplitude and pitch). • They can be described by their Pitch (P, in nm):• P= m x p, m-# of protein subunits per helical turn, p-axial rise

per subunit

12

VSV, a prototypical helical animal virus

• VSV (Vesicular stomatitis virus) coat protein: a “typical” helical coat protein.

• Small (50 aa derived from a 73 aa precursor), • alpha helical with 3 distinct domains• Domain characteristics are consistent with their function, • + charge interacts with nucleic acid, • hydrophobic with proteins on either side, • negative charge with polar environment. • Subunits are tilted 20o relative to the long axis of the

particle. P= 6.75 nm, m=4.5, and p=1.5.

• VSV Genome: 11,000 nt -ssRNA interacts with the nucleocapsid protein (N) to form a helical structure with P=5 nm.

• The particle is about 180 nm long and 80 nm wide.

Fig 4.4

13

14

15

Basic soccer ball and variations.Topology follows the constraints of Euclidean solid geometry.

5 3 2The icosahedron:12 vertices, 20 triangular faces, 30 edges.

5- , 3-, and 2-fold rotational axes. 5-fold rotational axes pass through the vertices.3-fold pass through centers of the triangular faces.2-fold pass through the edges.

Icosahedral structures

16

T=3

T=4

T=7l T=13l

Triangulation number (T)

•T=f2 x P where f=# of subdivisions on each side of a triangular face, P=h2 + hk + k2 where h and k are any nonnegative integer. •Only T’s that may be derived from the above equation are possible. •60 = minimal number of irregular subunits required •Beyond 60 subunits equivalence is not possible.

17

18

19

Picornaviridae, a prototype T=3 virus

• Fig. 4.9A illustrates quasi-equivalence with pentamer at each vertex and hexamers in other regions;

• Triangulation # = 3. • Note that VP-4 is not on the surface of the structure

but lies under the face.

20

Picornaviridae, a prototype T=3 virus • Fig. 4.9B. The protein subunits that form each protomer all

assume a similar (not identical) shape .• In fact all T=3 RNA viruses have proteins that form “8 strand

antiparallel barrels”. • The structures form from the polypeptide by first forming a

“jelly-roll barrel” that then goes on to form the wedge-shaped barrel when the capsid is being formed.

21

22

23

Complex structures

• Many, viruses are so large that they have evolved complex, more “cell like” structures.

• e.g. Poxviridae, paramyxoviridae, Coronaviridae)

Vaccinia virus, from Fig. 13.16

24

How do capsids interact with the genomic RNA rather than other

cellular RNA?

• In many cases viruses produce huge amounts of viral genome and inhibit cell mRNA synthesis (either directly or indirectly). This helps but does not solve the problem.

• Viral genomes have packaging signals (“psi”) that form structures recognized by one of the capsid proteins.

• Keep in mind that viruses never do anything correct 100% of the time (defective particles).

25

Where do virus capsids form?• Nonenveloped viruses

– Generally form in the cytoplasm (some in the nucleus) and mature before being released when the cell lyses.

• Enveloped Viruses-– Generally form the capsid as they are budding from a membrane. – Envelope proteins are inserted into the host cell membrane prior to budding. – Viral matrix and capsid proteins interact with the membrane and membrane

proteins at regions with a lot of envelope proteins. – The genome interacts with the nucleocapsid protein and budding is initiated. – Often the virion matures after release.

26From: http://home.ncifcrf.gov/hivdrp/Freed_figure.html