1. General Virology
VIRUS STRUCTURE

2. Virion vs virus Virion is the infectious particle
composed of nucleic acid, protein capsid, +/- envelope
may be extracellular or intracellular
Virus is any stage of infection

3. How do we know that NA is genetic material?
Hershey-Chase
Fraenkel-Conrat Experiment
TRANSFECTION EXPTS

4. TRANSFECTION FAILS FOR SOME VIRUSES
WHY?

5. Capsid Functions Protection of NA Attachment for naked viruses Enzyme
Helical vs Icosahedral Symmetry - Why do most viruses look alike?
Tobacco mosaic virus is a ssRNA virus composed of 6000 nucleotides. The capsid is made of 2100 copies of a single protein subunit that contain 158 amino acids. Calculate the percentage of the genome that is used for structure.

6. How do helical viruses differ?
Helical- one axis of symmetry down center
Multiple structural units

7. Icosahedral symmetry 20 identical equilateral triangles
Structural units on faces to give morphological capsomers
Pentons (5 fold axis of symmetry)
Hexons
3 fold through face
2 fold through edge
How do spherical viruses differ?

8. Envelope Attachment Entry Assembly- matrix proteins Release
Proteins are viral
Lipids are host
Rare in plants or bacteria - why?
If the membrane envelope is destroyed, the virus becomes noninfectious. Why?

9. Herpesvirus complexity
Tegument proteins - 12/84 viral proteins in HSV
Potential role?
Virion mRNA
DNAase virion nucleic acids
RT-PCR
probe genome array

10. Genome - DNA or RNA strandedness - (single) (double)
How do we experimentally show that DNA or RNA is the virus genetic material?
strandedness - (single) (double)
linear or circular, partial double stranded circle
number (single, segmented, multicomponent)

11. RNA Genomes sense (positive-sense, negative-sense, ambisense)
presence or absence of 5'-terminal cap or 5'-covalently-linked protein
presence or absence of 3'-terminal poly (A) tract
Retroviruses - replication strategy

12. Some viruses have high degree of secondary structure
Poliovirus - 5’ internal ribosome entry site (IRES)
Guest et al J. Virol. 78:

13. SARS/coronaviruses have conserved 3’region
SARS s2m in red
a - green = 530 loop of 16S RNA
Similar binding properties:
b - blue = S12
magenta = IF1
Possible role for s2m
Hijacks protein synthesis from cell(binding cell factors)
Needed to bind to similar viral protein for transcription
Potential drug target in red tunnel
Robertson et al PLOsBiology:3.

14. DNA Viruses may be large genomes
PolyDNAvirus (PDV) - contain many DNA segments
Mimivirus - larger than small bacteria

15. Host-induced modification
Viral property that varies depending on the host
Phage DNA hydroxymethyl cytosine (HMC) replaces C
Viral enzymes: C to HMC
Viral DNA polymerase: adds HMC not C
What is advantage of HMC?
Glucose is attached to HMC
Host enzyme needed to prepare glucose
Protects against host nuclease

16. What would happen if virus without glucose enters host with RE?
What would happen if virus with glucose enters host w/o enzyme to create UDP- glucose?
Host enzyme makes

17. Proteins structural proteins non-structural virion proteins
transcriptase,
protease
integrase

18. How to identify virion proteins
Purify KSHV virions
Run on SDS PAGE
Excise bands, digest - get sequence and compare to database

19. Chemical synthesis of poliovirus: What are the implications?
Small genome positive strand RNA - sequence known
Synthesized small DNA segments (~ 69 nucleotides) with overlapping complementary segments
Added a T7 phage promotor to DNA
Used DNA to make genome RNA in HeLa cell lysate with T7 polymerase
Results: How do you show success?

22. International Congress on Taxonomy of Viruses http://www. ncbi. nlm
Morphology
virion size
enveloped or naked nucleocapsid
capsid symmetry and structure
Genome characteristics
Replication strategy
Antigenic Properties

23. Baltimore classification

24. WHY TRANSFECTION FAILS

25. ONE STEP GROWTH CURVE 1939- Ellis and Delbruck:
Infection with a high multiplicity of infection (MOI): ratio of virus to host cell
Simultaneous infection
Single replication cycle
Sample at time intervals by plaque count for plaque-forming units (PFU),
Identification of latent phase
Determination of burst size/viral yield

26. Measuring Intracellular Events
Sample at time intervals after lysing cells ( Doermann)
Chloroform
Lysis from without
Identification of eclipse and maturation phases
Maturation phase

28. Strategy of replication
Lytic
Temperate

29. Biologic Properties natural host range mode of transmission in nature
vector relationships
geographic distribution
pathogenicity, association with disease
tissue tropisms, pathology, histopathology

30. How can you identify these viruses?