Download Microbial Genetics

Bacterial Genetics
Review
• Genome: genetic blueprint
• Gene:
• Most organisms-DNA
• Viruses
– DNA or RNA
Nucleic acids
• Nucleotides
– Sugar
– Phosphate
– Nitrogenous base
• DNA
– Guanine-G; cytosine-C; adenine-A; thymine-T
• RNA
– Uracil-U replaces thymine
Review
• DNA –ds helix
• Strands held together by H bonds
• Complimentary base pairing
– A pairs with T; in RNA, A pairs with U
– C with G
• Linear sequence of bases contains info
• 3 nucleotides code for 1 amino acid
Duplication of DNA
•
•
Replication of chromosome(s)
Precise duplication of DNA
•
DNA polymerase
•
Denaturation: separate strands
Terms
• Genotype: set of genes carry instructions
• Phenotype: expression of those genes
• Genome of prokaryotes includes
chromosomal DNA and plasmids
Flow of Information
• DNA to RNA to polypeptides
• Transcription-synthesis of complimentary
strand of RNA from DNA-mRNA
– Separates strands of DNA
– Copies only DNA needed for protein
synthesis-mRNA
Flow of Information
•
Translation: protein synthesis
– Decodes sequence of nucleotides into
amino acids (20)
– Genetic Code: codons, group of 3
nucleotides-AAU,CGC
•
AUG-start codon
Degenerate Code
• Each codon specifies a particular amino
acid
• Several codons code same aa
Flow of Information
• Translation
– Ribosomes-site of protein synthesis
– tRNA recognizes the specific codon
– tRNA forms complimentary base pairing with
the codon
– Has anticodon & carries the required aa
Question
• AUG is first codon on mRNA.
• What is the anticodon on the tRNA?
Regulation of Gene Expression
• Prokaryotes: Where is mRNA transcribed
in cell?
– Translation can begin before completion of
mRNA molecule
Regulation of Gene Expression
• Eukaryotes: Where?
– mRNA contains exons and introns
– Exons are expressed, encode for aa
– Introns do not
– Processing by ribozymes to remove introns &
splice exons
Regulation of Gene Expression
•
Conserve energy
•
•
•
Turn on & off transcription of genes
Constitutive genes not regulated
Operon
Operons
• Regulation of metabolic genes
• Uses repressors (regulatory genes)
– Block RNA polymerase from attaching
Induction
•
•
•
Turn on the transcription of gene
Inducer- induces transcription
Inducible enzymes
– Synthesized only when substrate is
present
• Glycolysis genes constitutive
Lac Operon
• Inducible operon: enzymes to metabolize
lactose
• Default is “off”
• Regulatory sites
– Promoter–
– Operator-
Lac Operon
• i genes code for repressor-regulatory
protein
– Always turned on
– Binds to operator
• Structural genes
– Lac operon-3 genes
Lactose in Medium
•
•
•
•
Binds repressor changing shape
Repressor can’t bind
RNA polymerase can bind
Enzymes for lactose metabolism produced
• To turn on operon:
Repressible Operon
•
•
Tryptophan operon
Usually occurs in anabolism
•
Repressor is inactive so tyrptophan is
synthesized
Default in “on”
•
Tryptophan Operon
• Amino acid in media
– Binds to repressor activating it
• Genes to synthesize amino acid produced
Mutation
•
Change (heritable) in base sequence of
DNA
–
–
–
Called mutant
Genotype differs from parent
Phenotype may be altered
Types of Mutations
• Spontaneous mutations
– DNA replication errors
– Occur at low frequencies
• Induced mutations-mutagens
– Alter structure of bases
– Errors in base paring
Point Mutations
• Change in 1 base-pair
– Single base is replaced with another
– Change in genotype
– May be change in phenotype
Types of Point Mutations
• Silent mutation- no phenotypic change
• Degenerate code
Missense Mutations
• Change in amino acid
• Can result in significant changes in
polypeptide
Examples of Missense
• Sickle cell anemia- hemoglobin
– Change from glutamic acid (hydrophilic) to
valine (hydrophobic)
• Change in shape of protein
Nonsense Mutation
•
•
Base-pair substitution
Create stop codon in middle of mRNA
– Premature termination of translation
Frame Shift Mutation
• Bases deleted or inserted
• Shifts translational reading frame
• Large insertions are transposons
Frame Shift
• THE FAT CAT ATE THE BAD RAT
• Remove a C
• THE FAT ATA TET HEB ADR AT
Induced Mutations
•
•
•
Mutagens
Increase mutation rate
Chemical mutagens
– Nitrous acid
– Occurs at random sites
Radiation
• UV light -nonionizing
• Covalent bonds between certain bases
• Adjacent thymines(Ts) can cross link
– Pyrimidine dimers
• Some enzymes that repair UV damage
Nucleoside Analog
• Structurally similar to normal bases
• Have altered base pairing
• 2 aminopurine replaces A & may pair with
C
• 5-bromouracil replaces T but may pair with
G
Genetic Recombination
•
Physical exchange of genes between 2
homologous DNA molecules
•
Contributes to population’s genetic
diversity
Horizontal Transfer
• Microbes of same generation
• Involves a donor cell - gives DNA to
recipient cell
• Part of DNA incorporated into recipient’s
DNA
Transformation
•
Free (naked) DNA in solution
– Cells after death, release DNA
•
Cells may take up DNA
– Only in certain stage of cell cycle
Competence
• Cells able to take up DNA & be
transformed
• Release competence factor that helps in
uptake
Transformation
• Enzymes cut DNA into small pieces
• Recombination between donor & recipient
• Few competent bacteria
First Evidence of Transformation
• Griffith in 1920s
• Streptococcus pneumoniae in mice
Conjugation
•
Mediated by one kind of plasmid
– F plasmid or F factor
•
Genes to control conjugation
•
Donor cells must have F plasmid
Differs from Transformation
• Cells must be of opposite mating types
– Donor is F plus
– Recipient is F minus
• Requires direct cell contact
• Transfers larger quantities of DNA
Conjugation
•
Gram negative cells
•
Gram positive cells produce sticky
surface molecules
– Keeps cells together
Conjugation
• Plasmid is replicated
– A copy of plasmid transferred to recipient
• F minus cell becomes F plus
• Receptors on new F plus change
F Plasmid
• Plasmid integrates into the chromosome
converts cell to Hfr cell( high frequency of
recombination)
• F factor DNA can separate and become
plasmid
Conjugation
• Hfr and F- cell
• Replication of Hfr begins in middle of
integrated F factor
• Small piece leads the chromosome into Fcell
• Donor DNA can recombine (DNA not
integrated is degraded)
Transduction
•
Bacterial DNA is transferred via a virus
-Bacteriophage
•
Virulent phages –lytic cycle
•
Generalized transduction
– Any gene on donor chromosome transferred
Generalized Transduction
• All genes are equally likely to be
packaged inside phage
– Virus cannot replicate in new bacteria
– Defective virus
• Specialized transduction-only certain
bacterial genes transferred
Plasmids
•
•
Extra chromosomal material
F factor or plasmid is a conjugative
plasmid
– Carries genes for sex pili and for transfer
Dissimilation Plasmids
• Enzymes break down unusual sugars and
hydrocarbons
– Pseudomonas use toluene and petroleum as
carbon and energy sources
– Used to clean up oil spills
– Allows bug to grow in adverse environments
Virulence Plasmids
• E. coli carries plasmids that code for
toxins –diarrhea
• Bacteriocins- toxic proteins kills other
bacteria
– E. coli produces colicins
Resistance Factors
• R factors- resistance to antibiotics, heavy
metals or cellular toxins
• Wide spread use of antibiotics- led to
selection of bacteria with R factors with
resistant genes
Transposition
• Segment of DNA moves from one place in
chromosome to another
– Rare event
– Transposable elements
– Insert within a gene & inactivate it
Transposons
• Contain information for own transposition
• Insertion sequences (IS) contain only
gene for transposase
• Complex transposons carry genes for
enterotoxins or antibiotic resistance