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Chapter 8
Microbial Genetics
part B
Genetics
• Study of what genes are.
• How DNA replicate.
DNA polymerase
DNA
DNA
RNA polymerase
• How information is expressed. Gene (DNA)
• Regulation of gene expression
Translation
mRNA
Protein
5’ ….ATGCCCTGAAAAGAGTCT…..3’
• How they carry information.
• Changes in the genetic material
• Genes and evolution
Regulation of Bacterial Gene Expression
• The cell conserves energy by making only those proteins needed
at a particular time.
– Constitutive enzymes are produced at a fixed rate
• Enzymes of glycolysis are example
• Genes encoded these enzymes – constitutive genes
– 60 – 80% of cell genes
–
Other enzymes are produced only as needed
• Inducible enzymes
– Induction – The process that turns on the transcription of a gene or
genes
• Repressive enzymes
– Repression – the regulatory mechanism that inhibits (turns off) gene
expression and decrease the synthesis of enzymes
Regulation of Bacterial Gene Expression
• Regulatory genes (I) code for regulatory proteins
– Often repressor proteins
• Repressor proteins then attach to a DNA segment known as the
operator (O).
– By binding to the operator, the repressor protein prevents the
RNA polymerase from creating messenger RNA.
– Blocking of gene expression is called repression
Repressor protein
Figure 8.13
Regulation of Bacterial Gene Expression
• Repressor protein activity depends on the presence or absence of an
effector substance.
Enzymes
Substrate
Product
• Induction – When the cells are exposed to the compound (substrate)
which has to be processed
• Repression - When the cells are exposed to a particular end-product.
Figure 8.13
Regulation of Gene Expression - Induction
I – Regulatory gene – encode repressor protein - active;
effector substance – substrate
Lac operon is called inducible operon
The lac operon is an operon required for the transport and metabolism of lactose in Escherichia coli and
some other enteric bacteria
Effector substance – Lactose
When Lactose is absent:
When Lactose is present into the cell:
- Repressor is active
- It is converted to allolactose (inducer)
- Repressor is inactive
- Active transcription – mRNA is synthesize
- No transcription
Induced by presence of Lactose (substrate)
Figure 8.14.2
Regulation of Gene Expression - Repression
I – Regulatory gene – encode repressor protein – inactive;
Effector substance – product
Tryptophan operon is repressible operon
Trp operon encodes the genes for the synthesis of tryptophan
Effector substance – Tryptophan
Absence of tryptophan
- Repressor is inactive
- Transcription is on
Presence of tryptophan
- Tryptophan (corepressor) binds with the
repressor protein – active repressor
- No transcription
Repressed by presence of Tryptophan ( product)
Figure 8.14.3
Regulation of Gene Expression - Catabolite repression
Lactose (disaccharide)  hydrolysis  glucose + galactose
Regulation of Lac operon also depend on the level of glucose in the medium
Glucose concentration in the medium controls the intracellular level of the small
molecule cyclic AMP (cAMP), when glucose is no longer available, cAMP
accumulates in the cell.
Glucose effect or catabolite repression – inhibition of the
metabolism of alternative carbon sources by glucose
Figure 8.15
Regulation of Gene Expression: Catabolite
repression
CAP – catabolite activator molecule
Lactose present
Glucose is absent
cAMP level is high
Lactose present
Glucose present
cAMP level is low
Transcription is stimulated
Transcription is not stimulated
Regulation of Gene Expression - Epigenetic Control
• Epigenetic control of gene expression - the switching on and
switching off of genes.
–
–
–
–
Methylating certain nucleotides – turned gene expression off
Methylated (off) genes are passed to offspring cells
Not permanent
Signals from the outside can work through the epigenome to change a
cell's gene expression
• Contributes to cellular differentiation and development
• Aberrant epigenetic control contributes to disease (particularly
to cancer)
• Biofilm behavior
Regulation of Gene Expression – Post-transcriptional control
• MicroRNAs control a wide range of activities in cells.
DNA
Transcription of
miRNA occurs.
miRNA
miRNA binds to target
mRNA that has at least six
complementary bases.
mRNA
mRNA is degraded.
Changes in the genetic material
1. Mutation
• A randomly derived change to the nucleotide sequence
of the genetic material of an organism.
2. Genetic Transfer and Recombination
• Genetic recombination refers to the exchange between
two DNA molecules.
– It results in new combinations of genes on the chromosome.
Mutation
• Mutation occur spontaneously – spontaneous mutation
• Mistakes of DNA polymerase
• Genetic recombination
– Mutation rate is the probability that a gene will mutate when a
cell divides; the rate is expressed as 10 to a negative power.
• Spontaneous mutation rate = 1 in 109 replicated base pairs
(frequency – 10-9 ) or 1 in 106 replicated genes (10-6 )
• Mutations usually occur randomly along a chromosome.
– A low rate of spontaneous mutations is beneficial in providing the
genetic diversity needed for evolution.
Mutation
• Based on the changes in DNA sequence mutations may be:
• Base substitution
• Deletion
• Insertion
• Based on the phenotypic effect on the organism mutations may be:
• Neutral
• Beneficial
• Harmful
• Organisms have mechanisms such as DNA repair to remove
mutations
• Mutation is generally accepted by biologists as the mechanism by
which natural selection acts
– Generating advantageous new traits – offspring survive and multiply
– Generating disadvantageous traits - offspring tend to die out
Base substitution (point mutation)
1.
Missense mutation - Change in one base result in change in amino
acid
2.
Nonsense mutation - Change in one base results in a nonsense
codon
Figure 8.17a, b
Mutation – deletion and insertion
• One or few nucleotide pairs are deleted or inserted in the DNA
• Frameshift mutation – shift the “translational reading frame”
Figure 8.17a, d
Identifying mutants - Replica Plating
• Positive (direct) selection
• Negative (indirect) selection
Example: an auxotrophic mutant cannot synthesize Histidine
100 colony
Figure 8.21
Mutagens
• Mutagens are agents in the environment that cause permanent
changes in DNA.
– Mutagens increase mutations to 10–5 or 10–3 per replicated gene
1. Chemical mutagens – Nitrous acid (alters Adenine), nucleoside
analogs, benzopyrene
2. Ionizing radiation (X rays and gamma rays) causes the formation of
ions that can react with nucleotides and the deoxyribose-phosphate
backbone.
3. UV radiation causes thymine dimers
– Light-repair separates thymine dimers
» Photolyases – enzymes that can
repair UV-induced damage
The Ames Test for Chemical Carcinogens
• Many mutagens have been found to be carcinogens
• Ames test –
– Mutated Salmonella his- (lost ability to synthesize histidine)
– Mutagenic substance may cause new mutation that reverse the original
mutation to his+ ( back mutation or reversions)
– Incubation with mutagen / Control – without mutagen
– Liver extract – supply all necessary activation enzymes
Figure 8.22
Genetic Transfer and Recombination
• Genetic recombination,
– The rearrangement of genes from separate groups of genes
– A molecule of nucleic acid is broken and then joined to a different one
– It contributes to genetic diversity.
• Vertical gene transfer occurs during sexual reproduction
– between generations of cells
•
Horizontal gene transfer in bacteria
– involves a portion of the cell’s DNA being transferred from
donor to recipient between cells of the same generation
Genetic Recombination
• In eukaryotes, recombination occurs in meiosis - chromosomal
crossover - exchange of genes between two DNA molecules
• Crossover occurs when two chromosomes break and rejoin
Figure 8.23
Genetic Recombination
• When some of the donor’s DNA has been integrated into the
recipient’s DNA, the resultant cell is called a recombinant
Donor
Recipient
Recombinant cell
Figure 8.25
Genetic transfer
1. Transformation
• During this process, genes are transferred from one bacterium to
another as “naked” DNA in solution.
• This process was first demonstrated in Streptococcus pneumoniae
and occurs naturally among a few genera of bacteria.
Figure 8.24
Genetic transfer
2. Conjugation
• Requires contact between living cells.
• Genetic donor cell is an F+;
–
–
F+ cells contain plasmids called F factors;
F factors are transferred to the F– cells during conjugation.
• Recipient cells are F–
– Converted to F+
Figure 8.27a
Conjugation
• When the plasmid becomes incorporated into the chromosome, the
cell is called an Hfr (high frequency of recombination) cell.
• During conjugation, an Hfr cell can transfer chromosomal DNA to
an F– cell.
• Usually, the Hfr chromosome breaks before it is fully transferred
Figure 8.27b
Plasmids
• Plasmids are self-replicating circular molecules of DNA carrying
genes that are not usually essential for the cell’s survival.
• Conjugative plasmids
– Carries genes for sex pili and transfer of the plasmid
• Dissimilation plasmids
– Encode enzymes for catabolism of unusual compounds
• Pathogenicity Plasmids
– Carrying genes for toxins or bacteriocins
• R factors
– Encode antibiotic resistance
• Plasmids usually occur naturally in bacteria, but are sometimes
found in eukaryotic organisms (e.g., the 2-micrometre-ring in
Saccharomyces cerevisiae)
Genetic transfer
3. Transduction by a Bacteriophage (viruses that infect
bacteria).
• In this process, DNA is passed from one bacterium to another in a
bacteriophage and is then incorporated
into the recipient’s DNA. .
2
In generalized transduction,
any bacterial genes can be
transferred.
Specialized transduction
3
4
5
6
Figure 8.28
Transposons
– Segments of DNA that can move from one region of DNA
to another
– Contain insertion sequences for cutting and resealing DNA
(transposase)
– Complex transposons carry other genes
Figure 8.30a, b
Phenotypic effect of mutation
• Neutral
• Beneficial
– Appear new quality
– Change metabolic requirement
• Loose the ability to metabolize some substrate
• Increase activity of some enzyme
• Harmful
• Lethal effect
Evolution
• Evolution is the process of change in the inherited traits of
a population of organisms from one generation to the next
.
1. Diversity is the precondition for evolution
• Genetic mutation and recombination provide a diversity of
organisms
2. The process of natural selection (positive
selection)
• Allows the growth of those best adapted to a given
environment
Learning objectives
• Explain the regulation of gene expression in bacteria by
induction, repression, and catabolite repression.
• Classify mutations by type, and describe how mutations are
prevented and repaired.
• Define mutagen
• Describe the effect of mutagens on the mutation rate.
• Compare the mechanisms of genetic recombination in bacteria.
• Differentiate between horizontal and vertical gene transfer.
• Describe the functions of plasmids and transposons.
• Outline methods of direct and indirect selection of mutants.
• Discuss how genetic mutation and recombination provide
material for natural selection to act on.