BACTERIOPHAGES

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BACTERIOPHAGES
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BACTERIOPHAGES - LYTIC GROWTH AND LYSOGENY PHAGE
STRUCTURE Capsid, Capsomer, chromosome Phage
T4 (Head, Collar, Tail, Core, Sheath, Base plate,
Spikes, Tail fibers, Specificity, Double
-stranded linear chromosome) LYTIC PHAGE
GROWTH Attachment (adsorption,
specificity) Penetration (injection) Replication
- Transcription, translation - Host provides
energy, ribosomes, RNA polymerase. low
molecular weight precursors for macromolecular
synthesis - Production of viral proteins and
nucleic acids Assembly (maturation) (packaging)
intact progeny viruses produced Lysis - release
of progeny Burst size Plaques (Host, Lawn,
Plaques) Phage growth in liquid cultures of
host Phages are said to infect their
host Phage preparations (i.e., suspensions of
phages in liquid) are also called phage
lysates TEMPERATE PHAGES AND LYSOGENY Lambda
- Infection Attachment, Penetration,
Circularization of chromosome. Repression of
lytic genes, Integration, Attachment site,
Lysogeny, Lysogenic immunity, Prophage,
Lysogen Prophage Induction Inducing agent
Repression abolished, Lytic gene
expression. Excision Lytic growth How would
you isolate a lysogenic culture if you are given
a temperate phage preparation and a culture of
host bacteria?
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PHAGE STRUCTURE
ICOSAHEDRON CHROMOSOME SINGLE-STRAND CIRCULAR
DNA 5,386 NUCLEOTIDES 10 GENES 30 nM TWENTY
TRIANGULAR PLATES NUCLEO - CAPSID
fX174
CAPSOMERS- STRUCTURAL PROTEIN
CAPSID
TMV - TOBACCO MOSAIC VIRUS
CHROMOSOME SINGLE-STRAND LINEAR RNA 6,000
NUCLEOTIDES CAPSOMERS INFECTIOUS ONLY RNA AND
PROTEIN CRYSTALS
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PHAGE T4
CHROMOSOME DOUBLE STRAND LINEAR DNA 2 x 105
NUCLEOTIDE PAIRS 1 x 108 MOLECULAR WEIGHT 200
GENES
HEAD
CAPSOMER
COLLAR
CORE
SHEATH
BASE PLATE
TAIL
TAIL FIBER (6)
EXTERIOR
SPIKES
INJECTION - PENETRATION
NOBEL HERSHEY
RECEPTOR PROTEIN
WALL - OUTER MEMBRANE
CYTOPLASMIC MEMBRANE
CYTOPLASM
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T4 GROWTH
MINUTES AFTER INJECTION
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STAGES

• 1. ADSORPTION / ATTACHMENT
• SPECIFICITY-RECEPTORS
• 2. INJECTION / PENETRATION
• SHEATH CONTRACTS
• CHROMOSOME INJECTION
• SYNTHESIS OF COMPONENTS (REPLICATION)
• TRANSCRIPTION
• TRANSLATION
• ENERGY
• PRECURSORS
• RIBOSOMES
• PRODUCES VIRAL mRNA
• VIRAL DNA (RNA) CHROMOSOME
• VIRAL STRUCTURAL PROTEINS

PROVIDED BY HOST
SUMMARY ONE PHAGE-INFECTED CELL PRODUCES 102
PROGENY IN ONE GROWTH CYCLE BURST SIZE AVERAGE
NUMBER PROGENY / INFECTED CELL
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PHAGE PLAQUES
107 HOST CELLS
TOP AGAR
AGAR PLATE
INCUBATE
CONFLUENT GROWTH
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PHAGE PLAQUES
107 HOST CELLS
10 T4 AND 107 HOST CELLS
TOP AGAR
TOP AGAR
AGAR PLATE
INCUBATE
CONFLUENT GROWTH
PLAQUES
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PLAQUE FORMATION BY LYTIC (VIRULENT) PHAGE
FREE PHAGES
30 MIN
MANY CYCLES
HOST CELLS INFECTED CELL
PLAQUE APPEARS CLEAR- HOST CELLS DESTROYED PHAGE
INVISIBLE
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GROWTH IN LIQUID CULTURE (OF HOST)
1 x 108 CELLS / ML
LOG TURBIDITY
TIME
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GROWTH IN LIQUID CULTURE (OF HOST)
1 x 108 CELLS / ML
LOG TURBIDITY
ADD 2-3 x 108 T4 / ML
PHAGE TITER?
TIME
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• ALL NUCLEIC ACID REPLICATION PROCEEDS THROUGH
  DOUBLE STRAND FORM
• T4 DOUBLE STRAND DNA DOUBLE STRAND PROGENY
  DNA
• MS2 SINGLE STRAND RNA CHROMOSOME

STRAND (ACTS AS mRNA) TRANSLATION YIELDS PHAGE
RNA REPLICASE (AND OTHER PROTEINS)





PROGENY
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BACTERIAL VIRUSES COMPARED TO BACTERIA
NO RIBOSOMES NO ENERGY GENERATING SYSTEM FEW
ENZYMES VIRULENT - LYSIS OF HOST TEMPERATE -
LYSIS / LYSOGENY
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REPRESSOR
PROPHAGE
BINARY FISSION
LYSOGENS LYSOGENIC PASSIVE REPLICATION OF
PROPHAGE DURING BINARY FISSION OF HOST
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TURBID PLAQUE FORMATION BY TEMPERATE PHAGE
FREE PHAGES
LYSOGENS
30 MIN
HOST CELLS INFECTED CELL
MANY CYCLES
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PHAGE LAMBDA - l - TEMPERATE LYTIC GROWTH OR
LYSOGENY 48,502 BP 30 GENES
COS
THE l CHROMOSOME
COS
COHESIVE SITE
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LAMBDA GROWTH ADSORPTION - PENETRATION
CHROMOSOME CIRCULARIZES LYTIC GROWTH LYSOGENY
5050
COS
COS
DNA LIGASE
COVALENTLY CLOSED CIRCLE
REPLICATION OR LYSOGENY
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TRANSCRIPTION OF REPRESSOR GENE
AND EARLY GENES TRANSLATION
PRODUCES REPRESSOR INTEGRASE
DNA REPLICATION REPRESSOR BINDS
OPERATORS INHIBITS TRANSCRIPTION OF
GENES IN LYTIC GROWTH
STIMULATES OWN TR
ANSCRIPTION COMPETITION REPRESSOR AND
LYTIC PROTEINS REPRESSOR WINS SHUTS OFF
LYTIC GENES INTEGRATION SITE SPECIFIC
RECOMBINATION BETWEEN

l DNA IS NOW PROPHAGE HOST IS NOW LYSOGEN
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l DNA INTEGRATION
ATTACHMENT SITE
HOST CHROMOSOME
REPRESSOR REPRESSION
GAL GALACTOSE BIO BIOTIN OPERON
INTEGRASE
PROPHAGE
LYSOGEN, STABLE, LYSOGENY PASSIVELY REPLICATED
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INTEGRATION DETAILS
5'
3'
HOST DNA
5'
3'
l DNA
INTEGRASE CUTS BOTH COMMON CORES
REJOINS l AND HOST DNA
l PROPHAGE
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PROPHAGE INDUCTION DNA DAMAGE REPRESSOR
CLEAVAGE LYTIC GENES NO LONGER INHIBITED
EXCISION, LYTIC GROWTH, PROGENY, LYSIS
REPRESSOR FRAGMENT
EXCISIONASE
REPLICATION
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LYSOGENIC IMMUNITY
l WILL NOT GROW ON A l LYSOGEN
PROPHAGE
REPRESSOR
l COMES FROM OUTSIDE AND INFECTS
NO l REPLICATION
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LYSIS / LYSOGENY DECISION

• A. ESTABLISHING LYSOGENY AFTER INFECTION
• REQUIRES cI cII, cIII, INT
• CRO ANTI-cI REPRESSOR (REQUIRED FOR LYTIC
  GROWTH)
• B. MAINTAINING LYSOGENY DURING HOST GROWTH
• REQUIRES cI

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C. CRO - BINDS OR3 - LIMITS TRANSCRIPTION
FROM PRM BINDS OR2 OR1 OL3 OL2 OL1 - TURNS
OFF EARLY FUNCTIONS D. cI - BINDS OR1 gt OR2
COOPERATIVELY INHIBITS LYTIC GENE
EXPRESSION CRO, cII, OP, etc STIMULATES
PRM MAINTENANCE cI STIMULATES ITS OWN
TRANSCRIPTION AND INHIBITS ALL OTHERS
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REGULATORY PROTEINS cI - REPRESSOR (FACILITATES
LYSOGENY) BINDS OR1 gt OR2 gt
OR3 BINDS OL1 gt OL2 gt OL3 cII - ACTIVATOR
OF PRE AND PI cIII - PREVENTS cII
DEGRADATION CRO - ANTI-cI REPRESSOR (FACILITATES
GROWTH) BINDS OR3 gt OR2 gt OR1 BINDS OL3
gt OL2 gt OL1
OPERATORS / PROMOTERS OR/OL - THREE OPERATORS
EACH PR AND PRM - OVERLAP OR PL -
OVERLAPS OL PRE - PROMOTER FOR REPRESSION
ESTABLISHMENT PRM - PROMOTER FOR
REPRESSION MAINTENANCE
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1. l DNA PENETRATES, CIRCULARIZED, HOST RNA
POLYMERASE TRANSCRIBES EARLY FUNCTIONS
2. cII ACTIVATOR STIMULATES TRANSCRIPTION FROM
PRE, PI
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3. CRO BINDS OR3 PREVENTS PRM FROM FUNCTIONING
4. cI REPRESSOR BINDS OR1 OR2 SHUTS OFF
EARLY GENES TURNS ON PRM
OL1 OL2
CRO / cI RATIO IS CRITICAL
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