GeneralVirology

VIRUSSTRUCTURE

 Virionvsvirus

Virionistheinfectious
particle
composedofnucleic
acid,proteincapsid,
+/envelope
maybeextracellularor
intracellular
Virusisanystageof
infection

 HowdoweknowthatNAisgenetic
material?

HersheyChase FraenkelConrat
Experiment

TRANSFECTION
EXPTS
TRANSFECTIONFAILSFOR
SOMEVIRUSES

WHY?

 Capsid
Functions
ProtectionofNA
Attachmentfornaked
viruses
Enzyme
HelicalvsIcosahedral
SymmetryWhydomost
viruseslookalike?
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.

 Howdohelicalvirusesdiffer?

Helicaloneaxisofsymmetry
downcenter
Multiplestructuralunits

 Icosahedralsymmetry

20identicalequilateraltriangles
Structuralunitsonfacesto
givemorphologicalcapsomers
Pentons(5foldaxisof
symmetry)
Hexons
3foldthroughface
2foldthroughedge

Howdospherical
virusesdiffer?

 Envelope
Attachment
Entry
Assemblymatrix
proteins
Release
Proteinsareviral
Lipidsarehost
Rareinplantsorbacteria
why?
If the membrane envelope is
destroyed, the virus becomes
noninfectious. Why?

 Herpesviruscomplexity

Tegumentproteins12/84viral
proteinsinHSV
Potentialrole?

VirionmRNA
DNAasevirionnucleic
acids
RTPCR
probegenomearray
Potentialrole?

 GenomeDNA
Genome or RNA

Howdowe strandedness - (single)

experimentallyshowthat (double)
DNAorRNAisthevirus
geneticmaterial? linear or circular,
partial double stranded
circle
number (single,
segmented,
multicomponent)

 RNAGenomes

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

 Someviruseshavehighdegreeofsecondary
structure

Poliovirus5internal
ribosomeentrysite(IRES)

Guestetal.2004.J.Virol.78:11097.

 SARS/coronaviruseshaveconserved3region

SARSs2minred
agreen=530loopof16SRNA
Similarbindingproperties:
bblue=S12
magenta=IF1
Possiblerolefors2m
Hijacksproteinsynthesisfrom
cell(bindingcellfactors)
Neededtobindtosimilarviral
proteinfortranscription
Potentialdrugtargetinredtunnel
Robertsonetal.2005.PLOsBiology:3.

DNAVirusesmaybelargegenomes

PolyDNAvirus(PDV)contain
manyDNAsegments

Mimiviruslargerthansmall
bacteria

 Hostinducedmodification
Viralpropertythatvaries
dependingonthehost
PhageDNAhydroxymethyl
cytosine(HMC)replacesC
Viralenzymes:CtoHMC
ViralDNApolymerase:adds
HMCnotC
WhatisadvantageofHMC?
GlucoseisattachedtoHMC
Hostenzymeneededto
prepareglucose
Protectsagainsthostnuclease

 Whatwouldhappenif
viruswithoutglucose
entershostwithRE?
Whatwouldhappenif
viruswithglucose
entershostw/o
enzymetocreate
Hostenzymemakes UDPglucose?

 Proteins

structuralproteins
nonstructuralvirion
proteins
transcriptase,
protease
integrase

Howtoidentifyvirionproteins

PurifyKSHVvirions
RunonSDSPAGE
Excisebands,digestget
sequenceandcompareto
database

Chemicalsynthesisofpoliovirus:Whatare
theimplications?
SmallgenomepositivestrandRNAsequenceknown
SynthesizedsmallDNAsegments(~69nucleotides)with
overlappingcomplementarysegments
AddedaT7phagepromotortoDNA
UsedDNAtomakegenomeRNAinHeLacelllysatewith
T7polymerase
Results:Howdoyoushowsuccess?

 QuickTime and a
TIFF (Uncompressed) decompressor
are needed to see this picture.


 InternationalCongressonTaxonomyofViruses
http://www.ncbi.nlm.nih.gov/ICTV/

Morphology Genomecharacteristics
virionsize Replicationstrategy
envelopedornaked Antigenic Properties
nucleocapsid
capsidsymmetryand
structure

 Baltimoreclassification

WHYTRANSFECTIONFAILS

 ONESTEPGROWTHCURVE

1939EllisandDelbruck:
Infectionwithahighmultiplicity
ofinfection(MOI):ratioofvirus
tohostcell
Simultaneousinfection
Singlereplicationcycle
Sampleattimeintervalsbyplaque
countforplaqueformingunits
(PFU),
Identificationoflatentphase
Determinationofburstsize/viral
yield

 MeasuringIntracellularEvents

Sampleattimeintervals
afterlysingcells(1952
Doermann)
Chloroform
Lysisfromwithout
Identificationofeclipse Maturationphase

andmaturationphases