1. Chapter 11 Lecture Outline
Viral Molecular Biology

2. Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.
Outline
11.1 Phage T4: The classic Molecular Model
11.4 A segmented (-) Strand RNA Virus: Influenza
11.5 A Retrovirus: Human Immunodeficiency Virus
Note: We will not cover 11.2, 11.3, 11.6!
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

3. Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.
Prokaryotic Viruses
Must bind to host cell receptor
Must cross a cell wall
In gram hosts 2 membranes to cross
Must not damage host cell initially
Use host nucleotides, amino acids, ATP
Replicate viral genome, build capsid, assemble new viruses
Exit through cell wall
Usually lyse host cells
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

4. Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.
Bacteriophage T4
Complicated structure
170 genes
10 different capsid protein types
Tail fibers bind host cell
Receptor = OmpC porin
Outer membrane protein
Long tail injects DNA
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

5. Phage T4 Adsorption and DNA Injection
Tail fibers bind to outer membrane (ompC, LPS).
Baseplate binds outer membrane
Tail sheath contracts, internal tube penetrates outer membrane
Injector digests peptidoglycan; internal tube penetrates cell wall
Injector contacts inner membrane; DNA expelled into cytoplasm.
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

6. Phage T4 Genome Replication
Upon entry, genome forms a circle
Early genes transcribed
Take control of cell, destroy cell chromosome
Replicates via Complicated structure
Uses cell nucleotides to replicate genome
“Rolling circle replication”
Continuous replication of many copies of genome
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

7. Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.
Rolling Circle
Concatemer
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

8. Phage Particles Self-Assemble
Late genes transcribed
Capsid particles
Head polymerizes around progeny DNA
Tail fibers, long tail made
Head, tail, tail fibers assemble
Lysis protein made
Destroys cell wall
Releases progeny
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

9. Summary Phage T4 Replication
The phage particle attaches to the outer membrane of E. coli and injects its genome.
Early genes are transcribed and translated into proteins, including nucleases to cleave host DNA and proteins for phage DNA replication.
Phage DNA undergoes rolling-circle replication, generating a multigenome concatemer.
Late genes are expressed to make head and tail components.
Phage genomes are packaged into heads.
Heads are assembled onto tails.
Tail fibers are added.
A phage-encoded lysozyme lyses the host cell, releasing about 200 completed phage particles.
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

10. Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.
Animal Viruses
Simpler entry into cell
Don’t have to cross a cell wall
Can enter through endocytosis
Or fusion of viral envelope to plasma membrane
More complex cycle in the cells
Must travel between organelles
Host transcription machinery in nucleus
Translation in cytoplasm
Transport via ER, Golgi, endosomes
Exit via cell lysis or budding
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

11. Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.
Viral mRNA
Prokaryotes: multiple proteins from 1 mRNA
Operon
Viral genome can be RNA or DNA
Multiple proteins created
Eukaryotes: 1 mRNA  1 protein
Viral DNA genome gives multiple mRNAs
RNA virus can create only 1 protein
Solutions:
1 protein cuts itself into smaller proteins (polio)
Multiple RNAs in a single virion (influenza)
RNA used to make DNA  multiple proteins (HIV)
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

12. (-) Strand RNA Virus: Influenza
Pandemic of 1918
Greatest one-year loss of life in recorded history
RNA inside shell of matrix proteins
Inside lipid envelope
2 major envelope proteins
Neuraminidase
Hemagglutinin
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

13. (-) Strand RNA Virus: Influenza
8 separate RNAs in genome
Each encodes one protein
(-) strand RNA can’t be read by ribosome
Must be transcribed to (+) RNA
No replicating RNA polymerase in host
All (-) RNA viruses must bring own polymerase protein into host cell
Influenza binds to host sialic acids
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

14. (-) Strand RNA Virus: Influenza
(-) strand RNA moves to nucleus
(+) strand mRNA synthesized
Move to cytoplasm
Viral proteins made
Envelope proteins placed in plasma membrane
(+) strand used to make progeny (-) RNA
Assembly at plasma membrane
Budding to release progeny viruses
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

15. (-) Strand RNA Virus: Influenza
Viral RNA polymerases are inaccurate
Introduce many mutations
Antigenic drift
Rapid evolution
New flu virus species every year
New vaccine necessary
Cell infected by 2 strains can recombine
Assemble new combinations of RNAs
Reassortment = antigenic shift
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

16. Human Immunodeficiency Virus
Bullet-shaped capsid
Encloses 2 identical copies of RNA
Plus polymerase proteins
Surrounded by envelope
Envelope proteins embedded
Binds to receptor
CD4 protein
Immune system T cells
Microglia cells in brain
AIDS-related dementia
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

17. Human Immunodeficiency Virus
Viral envelope fuses with plasma membrane
Genome released into cytoplasm
Polymerase replicates DNA from RNA
Reverse transcriptase
RNA destroyed as DNA made
Forms dsDNA
Acts as transposon
Moves to nucleus
Inserts into host genome
(+) ssRNA
(+) ssDNA
dsDNA
Integration into host genome
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

18. Human Immunodeficiency Virus
Reverse transcriptase is inaccurate
Introduces many mutations
Mutant viruses form within single individual
Single vaccine will not protect against all forms
5 major forms of HIV in humans
Many minor forms in each individual as disease progresses
From integrated DNA RNA is transcribed
RNA is used for progeny RNA, envelope and capsid proteins
Assembles in cytoplasm
Exits via budding
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

19. Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.
Retroviruses
Replicate transposon DNA
Insert repeatedly in chromosome
Destroy host cell
Eukaryotic cells have many retroelements
Old, inactivated retrovirus copies
Presence of many copies allows recombination
Movement of sections of chromosome
Moves pieces of genes together to make new genes
Major mechanism of evolution
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

20. Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.
Concept Quiz
Animal (-) strand RNA viruses always carry into the host cytoplasm their own:
genome and polymerase protein
genome and envelope proteins
genome and capsid proteins
Answer: A
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.

21. Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.
Concept Quiz
Why is it difficult to make a vaccine against HIV?
The virus mutates quickly inside an infected individual.
The envelope proteins cannot be isolated in the laboratory.
Different strains of HIV recombine to make new virus types.
Answer: A
Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc.