3.1 Infectious diseases bacterial genetics

central dogma

flow of genetic material DNA to RNA

explain bacterial plasmid uptake

aka transformation; bacteria acquire and incorporate foreign DNA (such as plasmids) from their environment

where does a promoter bind?

at the -35 or -10 positions of DNA sequence of upstream of the gene, recognized by RNA polymerase for binding too

define codon

set of three nucleotides in the mRNA that are read by ribosomes

what is the start codon?

methionine AUG

ribosome starts making protein here

what are the three stop codons?

UAA, UAG, UGA

ribosome falls off the message and protein synthesis is terminated

define mutation

any change in the DNA that makes it different than WT

define genotype

genetic makeup, DNA sequence

define phenotype

physical trait; look or behave

a change in (BLANK) can lead to a change in (BLANK)

genotype, phenotype

list the 8 bacterial phenotypes:

• motile

• nonmotile

• encapsulated

• acid producing

• Lac-

• smooth colonies

• yellow colonies

• rough colonies

explain Lac⁻ mutant

bacteria cannot digest lactose so lactose crosses the membrane through the lactose permease (coded by the lacY gene) and is then broken down by β-Galactosidase (coded by the lacZ gene) into glucose

both lac Z and lac Y genes (Z more upstream) are controlled by a single upstream promoter, lac promoter making them an operon

genotype of lac mutant?

lacY⁻

phenotype of lac mutant?

Lac⁻

can a Lac⁻ mutant grow on lactose as a carbon source?

no!

What gene can produce a Lac⁻ mutant?

mutations in lacZ or lacY gene! you can achieve the same phenotype with different genotypes

five main types of mutations:

• base sub

• deletion

• insertion

• frameshift

• null

define base substitution and the three categories:

a change from one nucleotide to another; silent, missense, nonsense

define silent mutation

change in codon that does NOT change the amino acid

change in DNA → change in mRNA → no change in protein

define missense mutation

change in codon so it encodes a different amino acid

change in DNA → change in mRNA → change in protein

define nonsense mutation

change codon so it becomes a stop codon

change in DNA and mRNA

define deletion mutation

removal of DNA (could be small or entire genes)

define insertion mutation

add piece of DNA to gene, many large insertions caused by transposable elements (viruses or mobile DNA sequences called transposons)

define frameshift mutations

deletion or addition of nucleotides not divisible by three

change in DNA → change in mRNA → change in all amino acids at and/or after point of mutation

null mutation

mutation completely eliminates gene function

in bacteria mutations may result in a change of ability to:

• utilize a carbons source, such as lactose

• synthesize an aa

• resist an Ab

how do mutations occur?

1. spontaneously (error in repli)

2. exposure to mutagenic agents

   1. UV light

   2. NaNO3 metabolized to HNO2 nitrous acid which deaminates bases altering their base pairing

      1. C is deamin → U, which bp w A and not G

      2. leads to U bp w A and therefor T bp w A

what divides faster bacteria or eukaryotes?

bacteria!

two pathways to acquire genetic changes?

• vertical gene transfer: parent to offspring

• horizontal gene transfer (HGT)

what are the three types of HGT?

transformation, transduction, conjugation

new DNA from outside cell, from other cells, direct cell-cell and maintained through VGT

define transformation

new genetic material acquired from other cells

competent (able to import DNA) bacterial cell acquired new genetic material from its enviro

historical explanation of transformation:

live smooth colonies (smooth/glistening bc of sugar capsule making them unable to be phagocytosed) kill mice. live rough colonies do not kill mice.

inject live rough and dead smooth, the live rough transform into live smooth as the uptake the free lysed contents of the dead smooth

explain pili transformation

bateria use pili to fish for and grab DNA

free DNA in enviro are sensitive to …

… DNase, an enzyme that digests DNA and inhibits transformation

imported DNA fragments from transformation are (BLANK) than a bacterial chromosome

SMALLER, 1-10 genes

define transduction

intro DNA from outside the cell

bacteriophage transfers genetic material from one bacterium to another

define bacteriophage

viruses that infect bacteria - virulent aka “lytic” growth cycle

define conjugation

DNA can be maintained and propagated in subsequent progeny through VGT

what are the two types of bacteriophages?

• lytic phages: repli inside the cell and subsequently kill (lyse) the host cell

• lysogenic/temperate phages: integrate their phage genome into the bacterial chromosome (as prophage)

explain lytic phage generalized transduction:

• during repli the host DNA degraded

• random piece of host chromosome can be packaged by phages

• any region of chromosome can be transferred

• phage particles carrying host DNA can subsequently infect another bacteria

the idea: integrate to get taken up and kill other bacteria

explain specialized transduction by lysogenic phages:

• region of bacteria chromosome directly adjacent to prophage may be accidentally packaged during excision

• only these specific regions can be transferred to another bacterium

differences btwn the types of HGT

feature	transformation	transduction	conjugation
Definition	Uptake of free DNA from the environment.	Transfer of DNA via a bacteriophage.	Direct transfer of DNA via cell-to-cell contact.
DNA Source	Free/naked DNA.	DNA packaged in a virus.	Plasmid or chromosomal DNA from donor.
Mediated by	Competence factors in bacteria.	Bacteriophages (viruses).	Sex pilus and conjugative plasmids.
Requirement	Competent bacterial cells.	Bacterial infection by a phage.	F+ or Hfr donor cell; direct contact.
Type of DNA	Plasmid or chromosomal fragments.	Bacterial genes or viral DNA.	Typically plasmid (e.g., F plasmid).
Example	Uptake of antibiotic resistance genes.	Phage-mediated gene transfer (e.g., λ).	Transfer of F plasmid in E. coli.

could the incoming DNA from transformation or transduction be maintained in the cell?

yes, there is homology

homologous recombination:

genetic process where two DNA molecules exchange genetic material by aligning and crossing over at regions of high sequence similarity

1. incoming DNA (made into ssDNA) lined up to homologous DNA region by RecA

2. double recombo result in acquisition of new gene in one strand of chromosome

3. recipient incorporate this new gene in one of the products of a round of chromosomal DNA repli (completely integrate; may get new ability, Ab resistance, virulence factor, new enzyme)

what are plasmids?

non chromosomal DNA

definition and types of replicons:

DNA elements capable of self-repli, chromosomes and plasmids

what two things do you need to be a replicon?

• ORI: site/sequence in DNA where rep protein binds

• rep gene: bind to ORI to initiate DNA repli

how do bacteria acquire new plasmids?

• bacteria can pick up free DNA plasmids by transformation

• some phages can transduce a whole plasmid

• plasmids can be transferred from one bacterium to another through conjugation

what is conjugation:

process by which genetic material is transferred from one bacterium to another through direct contact; and give donor ability to transfer plasmid to other bacterial cells (mating)

• unidirectional w donor and recipient

• DNA transferred as a plasmid (distinct from chromosome)

• plasmids carry special conjugation gene (tra gene) and are called conjugative plasmids

Conjugation summary

1. donor synthesizes conjugative or sex pilus, tra genes found on conjugative plasmid

2. donor sex pilus attach to recipient cell

3. upon contact, donor retracts sex pilus and pore formed btwn two cells, allowing transfer of ss copy of plasmid into recipient cell

4. ss copy of conjugative plasmid circularizes and repli completes complementary strand. upon acquiring this conjugative plasmid the recipient cell has become a new donor