Chapter 6
The Genetics of
Microorganisms
© 2004 Wadsworth – Thomson Learning
Structure of DNA
• Two strands
• Nucleotides
– Hydrogen bonds
between strands
– Neighboring
deoxyribose connected
• 3’ of one deoxyribose to
5’ of next deoxyribose
• Phosphate in between
• Double helix
• Base pairing
– G and C
– A and T
Figure 6.1
© 2004 Wadsworth – Thomson Learning
Roles of DNA
• Replication
– cell division
– need accurate copy
• Gene expression
– DNA
– RNA
– Protein
Figure 6.2
© 2004 Wadsworth – Thomson Learning
DNA Replication
• Semi-conservative
– old strand-template
– new strandcomplementary
• Replication fork
–
–
–
–
–
multiple enzymes
DNA unwinds
exposes nucleotides
synthesize new strand
one direction: 5’ to 3’
Figure 6.3
© 2004 Wadsworth – Thomson Learning
DNA Replication
• Complementary nucleotides match (A=T; G=C)
• DNA polymerase III binds nucleotides releasing
pyrophosphate
Figure 6.3
© 2004 Wadsworth – Thomson Learning
Bacterial chromosomes
• Replication of circular
chromosome
• Origin of replication
– bubble forms
– DNA unwinds
• Replication occurs in
both directions
• Two replication forks
• Continues until
replication forks meet
• Strands separate
Figure 6.4
© 2004 Wadsworth – Thomson Learning
Replication fork
Figure 6.5
© 2004 Wadsworth – Thomson Learning
DNA Replication
• Leading strand
– replication is continuous (5’ to 3’)
• primase makes primer
• DNA added to primer
• fork opens and replication continues
• Lagging strand
– polymerization in only one direction
• can’t go 3’ to 5’
– short segments synthesized (Okazaki fragments)
• when fork opens, new primer is made
• synthesis in direction away from fork
• fragments are joined together by DNA ligase
© 2004 Wadsworth – Thomson Learning
Transcription
• RNA polymerase
binds DNA at site of
promoter
Figure 6.6
© 2004 Wadsworth – Thomson Learning
Transcription
• DNA unwinds
• nucleotide bases are
exposed
Figure 6.6
© 2004 Wadsworth – Thomson Learning
Transcription
• ribonucleotides pair
with exposed bases
– uracil in RNA
replaces thymine
– U binds A
• ribonucleotides are
polymerized into
growing RNA chain
Figure 6.6
© 2004 Wadsworth – Thomson Learning
Transcription
• Termination
sequence
• Release of transcript
• single strand RNA
• DNA closes
Figure 6.6
© 2004 Wadsworth – Thomson Learning
Transcription
• Role of RNA from transcription
– mRNA
• template which encodes the protein
– tRNA
• transfer amino acids used to build the protein
– rRNA
• part of ribosome which is the site of protein
synthesis
• All used for translation
© 2004 Wadsworth – Thomson Learning
Translation
• Production of proteins
• Based on genetic
information of DNA
• Genetic code
– Codon has three
nucleotide
– Four different
nucleotides
– 64 possible
combinations
– 20 amino acids
• Redundancy
• Nonsense codons
Figure 6.8
© 2004 Wadsworth – Thomson Learning
Translation
• tRNA
– binds an amino acid
• specific amino acid for
each tRNA
– Anticodon
• recognizes codon
• three nucleotide
sequence in mRNA
which encodes a
specific amino acid
– activated with ATP
Figure 6.7
© 2004 Wadsworth – Thomson Learning
Translation
• Ribosome binds to
mRNA
– specific region
– start codon
• Methionine
– Ribosome binding
region
• Shine-Dalgarno
sequence
Figure 6.9
© 2004 Wadsworth – Thomson Learning
Translation
• tRNA with appropriate
anticodon and specific
amino acid binds to the
codon on the mRNA
– A site
• second tRNA binds in
similar fashion
– P site
• two amino acids are
joined in a peptide bond
Figure 6.9
© 2004 Wadsworth – Thomson Learning
Translation
• Ribosome moves
along mRNA
• first tRNA without
amino acid is
removed
• second tRNA with
both amino acids
moves to P site
Figure 6.9
© 2004 Wadsworth – Thomson Learning
Translation
• New tRNA enters A site
• Growing amino acid chain is transferred to
new amino acid
Figure 6.9
© 2004 Wadsworth – Thomson Learning
Translation
• steps repeat
– ribosome moves
– one codon at a time
• protein chain
– one amino acid
added for every
codon
Figure 6.9
© 2004 Wadsworth – Thomson Learning
Translation
• Continues until
nonsense (stop)
codon is reached
• no tRNA matches
• ribosome is removed
• protein chain is
released
Figure 6.9
© 2004 Wadsworth – Thomson Learning
Transcription and Translation
• Simultaneous
transcription and
translation
• mRNA chain is
transcribed
• translation begins
• multiple ribosomes on
single mRNA
– polysome
Figure 6.10
© 2004 Wadsworth – Thomson Learning
Regulation of genes
• Transcription
– Production of
regulatory proteins
• Bind DNA near the
promoter
• Example: Lactose
operon
– Interruption of
transcription
• Attenuation
• Translation
– Ribosomal proteins
• Global regulation
– Catabolite
repression
– Nitrogen regulation
– Phosphorus
regulation
– Stringent response
– Heat shock proteins
© 2004 Wadsworth – Thomson Learning
Transcriptional regulation
lac operon
• lacZ
• lacY
• lacA
– regulated by lacI
• Lactose absent
– repressor binds
– stops transcription
Figure 6.11
© 2004 Wadsworth – Thomson Learning
Transcriptional regulation
• Lactose present
– repressor bound
by product of
lactose
• allolactose
– transcription
occurs
– gene products of
all genes are
made
Figure 6.11
© 2004 Wadsworth – Thomson Learning
Attenuation
• Histidine operon
– Histidine present
– Leader protein made
• Translation occurring
simultaneously with
transcription
• Requires histidine
– Attenuator loop forms on
mRNA
• Displaces RNA polymerase
• Stops transcription
Figure 6.12a
© 2004 Wadsworth – Thomson Learning
Attenuation
• Histidine absent
– Leader protein not
made
• Not enough histidines to
complete protein
– Antiterminator loop
forms
• Prevents attenuator loop
from forming
• RNA polymerase
continues
Figure 6.12b
© 2004 Wadsworth – Thomson Learning
Regulation of translation
• Expression of ribosomal proteins
– Unused proteins bind to encoding mRNA
– Inhibit translation
Figure 6.13
© 2004 Wadsworth – Thomson Learning
Two component regulation
• Phosphorylation of
sensor
• Phosphate passed to
response regulator
• Response regulator
reacts with DNA
changing gene
expression
– Increase
– decrease
Figure 6.14
© 2004 Wadsworth – Thomson Learning
Genetic Information
• Genome
– total DNA of a cell
– most have single circular chromosome
– some have linear chromosome
• Plasmids
– small, circular, extrachromosomal DNA
• encode beneficial factors
• resistance factors (antibiotic)
• conjugative plasmids
– transfer to other cells
• Genotype: genetic makeup
• Phenotype: appearance and function
© 2004 Wadsworth – Thomson Learning
Changes in Genetic Information
• Mutations
– chemical change
in DNA
• chemical
mutagens
– Bind DNA
– Change in DNA
• physical mutagens
– UV light
– Ionizing radiation
• biological mutagens
– Transposable
elements
» Insertion
sequences
» transposons
Figure 6.16
© 2004 Wadsworth – Thomson Learning
Consequence of mutations
• Types of mutations
– base substitution
• wrong nucleotide
– deletion mutation
• Results
– Lethal mutation
– Conditional expressed
mutations
• nucleotides deleted
– Inversion
• reverses order of a
segment
– Transposition
• moves a segment of
DNA
– Duplication
• identical new segment
Figure 6.15
© 2004 Wadsworth – Thomson Learning
Physical mutagen--UV damage
• UV light
– stimulates neighboring
bases to form dimers
• thymine dimers
– activate repair
systems
Figure 6.17
© 2004 Wadsworth – Thomson Learning
Physical mutagen--UV damage
• Thymine dimers
distort the DNA
structure
• Enzymes remove
the damaged
nucleotides
Figure 6.17
© 2004 Wadsworth – Thomson Learning
Physical mutagen--UV damage
• Repairs may result
in incorrect
nucleotide
replacement
• Mutation is result
Figure 6.17
© 2004 Wadsworth – Thomson Learning
Selecting and identifying mutants
• Direct selection
– Conditions favor growth of desired mutant
– Growth of bacteria in presence of antibiotic
– Only successful growth are mutants
• Indirect selection
–
–
–
–
Prevent growth of mutant
Kill growing cells
Desired mutants larger percentage of population
Isolate mutants
• Site-directed mutagenesis
– Recombinant DNA manipulation
© 2004 Wadsworth – Thomson Learning
Selecting and identifying mutants
• Brute strength
– Screen large numbers
– Replica plating
• Transfer large numbers of colonies
• Track growth
Figure 6.18
© 2004 Wadsworth – Thomson Learning
Ames Test
Figure 6.19
© 2004 Wadsworth – Thomson Learning
Transformation
• DNA exits one cell, taken up by another cell
– Natural
• few bacteria take up DNA fragments
– Artificial--induced in laboratory
• useful tool for recombinant DNA technology
Figure 6.20
© 2004 Wadsworth – Thomson Learning
Conjugation
• Conjugative plasmids
–
–
–
–
plasmids transfer
genetically encoded
F plasmid in E. coli
sex pilus connect two
cells
• one cell F+
• one cell FFigure 6.21
© 2004 Wadsworth – Thomson Learning
Conjugation
– One strand of
plasmid DNA is
broken (nicked)
– replication begins
– synthesized linear
strand enters F- cell
– linear strand is
copied forming a
complete plasmid
– both cells are F+
Figure 6.21
© 2004 Wadsworth – Thomson Learning
Transduction
• Bacteriophage
– virus that infects
bacteria
– reproduce in bacteria
– some phages contain
bacterial DNA
• rare event
• transducing particle
– cell lysis and release
• normal phage
• transducing particles
Figure 6.23
© 2004 Wadsworth – Thomson Learning
Transduction
– Transducing particles
• infect other bacteria
• inject bacterial DNA into
new cell
– genetic exchange
• one bacteria cell to
another
– integration into
chromosome
Figure 6.23
© 2004 Wadsworth – Thomson Learning
Eukaryotic Microorganisms
• Genetic
exchange
– Similar to plants
and animals
– Haploid
gametes fuse
Figure 6.23
© 2004 Wadsworth – Thomson Learning