Chapter 7
Microbial Genetics
The Blueprint of Life: from DNA to
Protein
The Central Dogma of Molecular
Biology
• The flow of ______to _______to ________
– DNA is _______________into RNA
– RNA is _______________into Protein
5´
3´
3´
5´
DNA
(genotype)
Transcription
5´
Translation
by ribosomes
3´
mRNA
Polypeptide
NH2
Phenotype
Overview
• The world contains a vast variety of organisms
• Every trait or characteristic of each organism
from shape to function is determined by the
information within its DNA
– 4 coding units
•
•
•
•
Adenine
Thymine
Cytosine
Guanine
Genome
• The complete set of genetic information
within a cell
–
• Includes:
• Composed of DNA
• Some viruses have RNA genomes
– Retroviruses such as HIV
Chromosome
• The E. coli chromosome is 4.7 x 10e6 base
pairs (bp) long
• The size of an average bacterial gene is 1000
bp
• E. coli has about 3,500 genes
• Humans have about 30,000
Plasmid
• Plasmids
– Have genes for their own replication
– May confer new characteristics to the cell:
• ____________________(R plasmids)
• ____________________(bacteriocin toxin which kills
bacterial species that lack the plasmid)
• ____________________(genes enabling a bacterium to
become pathogenic)
– May contain genes that mediate plasmid transfer to
new cells
• ____________________(F plasmids)
Gene
• The functional unit of the genome
• Encodes a product
– Transcribed to RNA
– Translated to a protein
– Some genes only go to RNA
Characteristics of DNA
• Deoxyribonucleic acid
– ______________attached to __________(pentose
sugar)
– Attached to one of 4 nitrogenous bases
• A,T,C,G
• Joined by a ____________bond between a 5`
phosphate and a 3` hydroxyl
– The designations 5` and 3` refer to the numbered
carbon atoms of the pentose sugar of the
nucleotide
Characteristics of DNA
• The joining of nucleotides forms a series of
alternating sugar and phosphate units
– Sugar-phosphate backbone
Characteristics of DNA
• DNA always has a 5` phosphate at one end
and a 3` hydroxyl at the other
– 5` end and 3` end
• DNA in a cell is ____________and ________in
structure
Characteristics of DNA
• The 2 strands of double-stranded DNA and
have ______________________________
– ____________bonds with 2 hydrogen bonds to
_____________
– _____________ bonds with 3 hydrogen bonds to
_____________
Characteristics of DNA
• The 2 strands of DNA are also _____________
– Oriented in opposite directions
– 5` to 3`
– 3` to 5`
Characteristics of RNA
• Comparable to DNA in many ways
• Exceptions:
– RNA is made of ____________________not
deoxyribonucleotides
– RNA has _____________ instead of ___________
– RNA is usually ____________stranded
– RNA is usually ____________not helical
– RNA is found in shorter strands
Characteristics of RNA
• 3 functional groups of RNA
– Messenger RNA (mRNA)
– Ribosomal RNA (rRNA)
– Transfer RNA (tRNA)
Characteristics of RNA
• Transcription or making RNA
– __________is used as a template
– Most genes (DNA) code for proteins
– The genes need to be ____________into ______________
________________
– The mRNA is then ______________into __________during
protein synthesis
5´
3´
5´
3´
DNA
(genotype)
Transcription
5´
3´
mRNA
Characteristics of RNA
• _______and ________are not translated into
protein
• They are the ultimate products
• Each are involved in protein synthesis
Bacterial DNA replication
• Before a cell divides DNA must be replicated
creating a second DNA molecule
• Each cell after the division will have the _____
____________________
– The offspring are an _____________of the parent
cell
Cell replicates its DNA.
Binary
fissionmethod used
by most
bacteria for
reproduction
Nucleoid
The cytoplasmic
membrane elongates,
separating DNA
molecules.
Cross wall forms;
membrane
invaginates.
Cross wall forms
completely.
Daughter cells
may separate.
Cell wall
Cytoplasmic
membrane
Replicated
DNA
Bacterial DNA replication
• ____________________process that requires
monomers and ________to form the covalent
bonds
• Triphosphate deoxyribonucleotides serve both
functions
Bacterial DNA replication
• DNA replication is generally _______________
– Replication proceeds in _____________directions
– Allows for the entire chromosome to be replicated
in half the time
Bacterial DNA replication
• Replication is ____________________
– Each of the _________molecules contain ____of
the __________strands
• Template strand and synthesized strand
Original DNA
First replication
New strands
Original strands
Second replication
Bacterial DNA replication
• DNA replication requires a coordinated action
of many different enzymes and other proteins
•
•
•
•
•
•
Enzymes that synthesize new DNA strands
One strand is used as a template
Add nucleotides one at a time
Always ____________________________
New strand elongates in the ___________direction
Bacterial DNA replication
• The replication process is very accurate
– One mistake occurs every 10 billion nucleotides
– DNA polymerase has a “______________” ability
Bacterial DNA replication
The Process
1. Specific proteins recognize a region on the
chromosome : _______________________
2. _______________the double-stranded
molecule
3. _________synthesizes small fragments of
_________called ____________
Bacterial DNA replication
The Process
Bacterial DNA replication
The Process
4. Synthesis continuously proceeds of one new strand in
the 5` to 3` direction this is the ________________
5. ________________DNA polymerase cannot add
nucleotides to the ____________
–
–
Synthesis must be reinitiated periodically as the helicase
unwinds the DNA
Each re-initiation is preceded by the synthesis of an RNA
primer called an Okazaki fragment
6. __________seals the gaps between fragments on the
lagging strand
Gene Expression in Bacteria
• _____________: The properties of the cell
determined by the expression of the genotype
(gene expression)
• _____________the sequence of nucleotides in
DNA (genes)
• Gene expression involves 2 separate processes
–
–
Transcription
• DNA to RNA
•
– Transcription begins when _________________recognizes
a nucleotide sequence on DNA called a _____________
•
– Can initiate synthesis without a primer
– Once the first RNA polymerase is out of the way a second
RNA polymerase can bind initiating a new round of
transcription
• A single gene can be transcribed multiple times very fast
•
– Transcription ends when a RNA polymerase encounters a
terminator
Translation
• RNA to Protein
– This process requires _______________________
___________
• mRNA is a temporary copy of genetic
information
– The code is deciphered using the _________which
correlates sets of ___________________with one
amino acid
Translation
• Codons
– There are ____ codons
– ____ translate to the ______ different amino acids
– More than 1 codon can code for a specific amino
acid
The Genetic Code
Translation
• Ribosomes
– Made of ______________________
– The translation site
– Bring each amino acid into a favorable position so
an enzyme can catalyze a _______________
Translation
• tRNA
– Segments of RNA that ______________________
– Act as a key that interprets the genetic code
– The recognition is possible since the tRNA has an
________________
Translation
• Initiation
– Begins as the mRNA is being synthesized
– The first ______________starts the process
• Determines the ________________
Translation
• Elongation
– tRNA, is required:
• The amino acid is attached to one end of the tRNA
molecule
• The other end of the tRNA molecule carries a triplet of
bases, the anticodon, that is complementary to the
codon
Translation
• Termination
– Elongation stops when the ribosome reaches a
_____________
– The stop codon does not code for an amino acid
– Enzymes free the protein
Examples
• DNA: what is the complementary strand?
– 5` ACT TAG GAC AAA TAT CAT 3`
• Transcription: What is the mRNA sequence?
– 5` ATT TAC GCA GAC CCC AAT 3`
• Translation: What is the anticodon?
– AUG UUU CAU GAC AUU CAU
• What is the polypeptide of this mRNA sequence?
– 5` AUG AAA
GAU
UAU
UAU
UAA 3`
Regulation of Bacterial Gene
Expression
• Control mechanisms due to changes in the
environment
– Cells control metabolic pathways by 2 general
mechanisms
•
• Or the more energy efficient strategy: _____________
_________________, making only what is required
– To do this the cell controls the expression of certain genes
– Turns them off
Lac Operon model
• Operon: a set of genes controlled by a single
regulatory protein
• The _____________consists of 3 genes
involved with _________________in bacteria
– The effect is the genes are only expressed when
lactose is present but glucose is absent
Sensing and Responding to
Environmental Fluctuations
• Microorganisms adapt to changes in their
environment by altering the level of
expression of certain genes
– Chemical “sensing”
Quorum sensing
• “Sense” the density of cells within their own
population
• Activates genes that are only beneficial when
expressed in ___________________
– Cooperative activities which form biofilms are
controlled by this
Staphylococcus aureus
• Gram +, coccus
• Commonly called staph
• Responsible for skin infections
– Boils and pimples
– Surgical wounds
• Treated with methicillin (penicillin-like) since the 1970s
• Now 60% of S. aureus strains isolated are resistant to
methicillin
• Vancomycin considered the drug of last resort is now
showing to be ineffective
– 2002 diabetic foot ulcer, isolated a strain in our own Detroit
• This organism was also resistant to other common antibiotics,
including penicillin, methicillin, and ciprofloxacin
• How do multiple resistant strains arise and
evolve?
• How are resistance traits transferred readily to
other bacteria?
• Why must bacteria rely on these methods?
Genetic change in Bacteria
• All organisms need to adapt in order to
survive or succeed.
– Changing environmental conditions
• If you fail to succeed then other competing
organisms more “fit” will predominate
– Natural Selection
Genetic change in Bacteria
• Bacteria have 2 ways in which they can adjust
to changing conditions
–
–
Genetic change in Bacteria
• Genetic change alters the ____________of an
organism
• A change in genotype can easily alter the
physical or observable characteristics of an
organism: _____________
Genetic change in Bacteria
• Genetic change can occur by 2 mechanisms
–
–
Genetic change in Bacteria
How do multiple resistant strains arise and evolve?
• Mutation is a __________________________
_________________________
– Passed to daughter cells
– Mutant (modified organism)
• Since the mutation arises in a single cell then
passes it to its off spring
– _____________gene transfer
Genetic change in Bacteria
How are resistance traits transferred readily to other bacteria?
• Gene transfer occurs when a ______________
_____________________
– _____________gene transfer
Spontaneous Mutations
• Occur in a cells natural environment
– Random events
– 1 in 10 billion base pairs during DNA replication
Types of Mutation
• Change in DNA sequence
– Point Mutations
•
•
•
Also called Frameshift mutations
Substitution
• Suppose DNA code is represented by the letters:
• THECATATEELK
– Group them into triplets (like codons)
• THE CAT ATE ELK
• The substitution of a single letter or nucleotide
could change the meaning of the sentence or
result in a meaningless phrase
• THE RAT ATE ELK
• THE RAT RTE ELK
Insertion or deletion
• More serious changes
– Results in ______________
• Affect proteins much more seriously
• Why?
• THE CAT ATE ELK
• TRH ECA TAT EEL K
• TEC ATA TEE LK
Insertion
Deletion
Effects of Mutations on proteins
• _____________
– Does not change the amino acid sequence due to
redundancy of genetic code
• AAA changed to AAG (DNA)
• UUU changed to UUC (mRNA)
• Both codons are for phenylalanine= no change in
phenotype
Effects of Mutations on protein
• _______________mutation
–
– Effect depends on where in a protein it occurs
• If in critical location such as an active site of an enzyme
– Loss of function
• If in a less important location
– No adverse effects
Effects of Mutations on protein
•
– When a base pair substitution changes an amino
acid codon into a stop codon
• Results in non functional proteins
• Frameshifts (insertions or deletions)
– Result in drastic missense and nonsense
mutations
– Except when close to the end of a gene
Types of Mutations
Base Substitution Mutation
Missense Mutation- the new codon
results in a different amino acid
Base Substitution Mutation
Nonsense Mutation- new codon is a
stop, resulting in a shortened protein
Gene transfer
• Genetic Transfer -
results in Genetic Variation
– Genetic Variation - is needed for Evolution
• 3 Ways Genetic Transfer can occur in Bacteria
– 1.
– 2.
– 3.
• These methods are responsible for the spread
of antibiotic resistant genes!
DNA mediated Transformation
•
– Sources: lysed cells in a population of bacteria
– Some genera of bacteria secrete segments of DNA
Fredrick Griffith experiment 1928
• 2 Strains of Streptococcus pneumoniae
– 1. Virulent strain with a capsule - Pneumonia
– 2. Avirulent strain without a capsule - no disease
Non pathogenic strain
DNA from heat
killed virulent
strain
Transformed bacteria is now pathogenic
Conjugation
•
Transduction
•
• Bacteriophage
– virus that only infects bacteria
• How do multiple resistant strains arise and
evolve?
• How are resistance traits transferred readily to
other bacteria?
• Why must bacteria rely on these methods for
adaptation and evolution?