Download Microbial Genetics

Microbial Genetics:
From Genotype to Phenotype
Nucleic acid synthesis
Protein Synthesis
Heredity
Genetic Recombination
Microevolution
Microbial Genetics
Notable differences in hereditary
material in viruses, prokaryotes
and eukaryotes
Important basis for modern
methods of identification and
disease diagnosis
Review
Nitrogenous base
dNTP
Base pairing
DNA
RNA
Coding strand
Template strand
What does DNA do?
Bacterial cell
DNA
transcription
RNA
translation
DNA replication: Cell
Division requires
duplication of DNA
protein
Large molecules of DNA are called
chromosomes
Bacteria have circular chromosomes and
sometimes additional small, circular
pieces of DNA called plasmids
Clones
Replication of DNA
• Because DNA is double stranded replication can occur
on each strand simultaneously in opposite directions
• Continuous, newly synthesized strand added in 5’ to 3’ is
leading strand
• DNA synthesized from behind by the addition of Okazaki
fragments is lagging strand, olignucleotides are still
added to the 3’ end
DNA replication
Original double stranded DNA
Two new strands (green dotted lines) are created using a
polymerase enzyme
The process of creating the new strands is called polymerization
DNA polymerization
Sugar phosphate backbone
Nitrogenous bases
3’TAGCTTGCCTCTGAATGAGAATATGGCACCATCGAAA…………….5’
5’ATCGAACGGAGACTTACTCTTA3’
DNA
Polymerase
G
T
A
A
A
C A
G
T
C
T
A
dNTPs are are added to the new strand
(green) so that the dNTPs pair with the
corresponding ones on the complimentary
strand (black)
DNA Polymerization
• Polymerization, requires DNA polymerase enzyme
• Synthesis occurs only from 5’ to 3’ of new strand
• dNTP incorporated into 3’ end of new strand by DNA
polymerase
• Formula for polymerization:
(dNMP)nDNA +dNTP(dNMP)n+1DNA + PPi
• (dNMP)nDNA is the growing strand
• dNTP is the deoxynucleotide triphosphate
• (dNMP)n+1DNA is the growing strand after a dNTP is
added
• iPP is inorganic diphosphate
Original strands are separated generating replication forks
5’
3’
Synthesis occurs on both
strands of original DNA in
opposite directions
Leading strand
Okazaki
fragments
Lagging strands
Leading strand
Nucleic acid polymerization
Nitrogenous base
5’
Nitrogenous base
Nitrogenous base
Nitrogenous base
Nitrogenous base
Nitrogenous base
Polymerase
enzyme
Nucleotide triphosphate
Nitrogenous base
Inorganic
phosphate
Transcription is synthesis of RNA from DNA
• NTPs (ATP, CTP, GTP, UTP) incorporated in to strand of
mRNA (RNA polymerization)
• mRNA is complementary to the template strand of DNA
• RNA polymerase enzyme is used
What is a gene?
tRNA
rRNA
DNA
mRNA
protein
Synthesis of mRNA
Double stranded DNA has a coding strand (+) and template strand (-)
5’
3’
+
Original strands are separated where transcription is to begin
mRNA synthesis occurs on the template strand (-) of original DNA
Synthesis of mRNA
Template strand of DNA
3’TACCTTGCCTCTGAATGAGAATATGGCACCATCGAAA…………….5’
5’AUGGAACGGAGACUUACUCUUA3’
RNA
Polymerase
G
U
A
A
A
C A
C
U
AUG is start codon
A
G
U
Genetic Code for Building Proteins
Every three bases (triplet) of DNA corresponds to a
codon of mRNA which corresponds to an anticodon in
tRNA which bares a specific amino acid
Use the table in your book to provide the amino acids for
the following DNA sequence on the coding strand :
CGTCCCGTC
Eukaryotes
E I
E I
E I
E I
E I
E I
E I
E
DNA contains introns and exons
I
I
I
I
I
I
E
E
E
E
E
I
E
Exons have the code to build proteins
E
E
Components of Prokaryote Ribosome
21 different proteins
16SrRNA
30S subunit
31 different proteins
70S ribosome
(prokaryotes)
50S subunit
23S rRNA and
5S rRNA
Ribosomes
The site of protein synthesis
Prokaryotes have 70S ribosomes
Eukaryotes have 80S ribosomes
Because ribosomes do such an important job
drugs that inhibit them have drastic effects on the
cell (many antibiotics and the toxin ricin inhibit
protein synthesis by binding to ribosomes)
The genes for ribosomal RNA are often used to
measure the similarity between organisms
16SrRNA gene for bacterial systematics
18SrRNA gene for eukaryote systematics
Translation
Growing polypeptide
connected by
enzymatic linkage
between amino acids
tRNA
mRNA
ribosome
Example of regulation of protein synthesis:
Enzyme Induction
Prom. Oper.
Gene
Pol.
Substrate binds to repressor allowing
polymerase to transcribe mRNA
repressor
mRNA
substrate
Enzyme acts on
substrate
Enzyme
synthesized
Example of regulation of protein synthesis:
Enzyme Induction
Prom. Oper.
Pol.
repressor
Gene
Gene turned ‘off’
In the absence of
substrate, the
repressor blocks
polymerase enzyme
Note: the status of a gene being turned on or
off is not heritable. While it allows individual
organisms to adapt to changing
circumstances, it does not effect the evolution
of species as does mutation.
Mutation
• Point mutation -results from the replacement of a
nitrogenous base effects may 1. be “silent” causing no
change in the protein structure, or 2. result in altered
protein with different performance or 3. produce protein
that doesn’t work
• Frameshift mutation -results from the insertion or
deletion of bases, has dramatic effects on protein, often
deleterious.
The genetic code is analogous to written
language
Suppose the sentence below is a genetic
sequence:
the red fox ran out
After a point mutation the sentence might be:
the red fax ran out
After a frame shift mutation (deletion of letter e)
the sentence might be:
thr edf oxr ano ut
Mutagens
• Chemicals that bind to DNA and affect the process of
replication or transcription. Some naturally occurring
chemicals are mutagenic, many industrial chemicals are
mutagenic
• Radiation, including UV light, is mutagenic
Genetic Variation Results from Mutation
Most mutations are either harmful, or neutral, but
sometimes they are beneficial.
If the mutations are not too harmful, they will be
passed on to their progeny (offspring). This is the
hereditary basis of evolution.
These heritable changes in a lineage or populations of
organisms over generations contribute to microevolution
Antibiotic Resistance in Bacteria
• Bacteria can multiply rapidly into large populations that
reach a stationary phase
• Antibiotic therapy or accidental ingestion of antibiotics
acts as an agent for natural selection
• Single point mutation in DNA can lead to resistance in a
single mutant bacterium
• Mutant ‘lost in crowd’ until antibiotic therapy kills off
susceptible bacteria
• Mutant becomes dominant in population and gives rise
to mutant clones
Variation by Mutation is Compounded by
Genetic Recombination
•
•
•
•
•
Sexual reproduction
Bacterial transformation
Bacterial conjugation
Virus-mediated gene transfer
Other transfer between symbionts
Sexual reproduction is genetic
recombination
parent cells
produced
gametes by
meiosis
Gametes
fuse to form
a zygote
zygote
gametes
2N diploid
stage
gametogenesis
fusion of
gametes
1N haploid
stage
egg
sperm
Bacterial Transformation
Heat-killed bacterium
disintegrate and release
DNA into surroundings
Bacterium with gene for
capsule
Bacterium without gene
for capsule
Genes taken in
and expressed
Conjugation
1.
2.
Plasmids may contain genes that allow a bridge (pilus)
to form between two bacteria. Bacteria with these
plasmids are F+
The plasmid from the F+ (donor) bacterium is
replicated and transferred to the F- (recipient)
bacterium
pilus
F+
1.
F+
F-
2.
F-
High frequency recombinations
chromosome
Plasmid inserts into
chromosome
F+
recipient
Hfr+
Hfr+
recipient
Hfr+ DNA transferred
into recipient and
inserted into
chromosome
Transposable elements (jumping genes)
• A mobile genetic sequence that can move from one
plasmid sequence to another sequence or to a
chromosome
• May result in the disruption of gene activity
• Make up a large portion of Eukaryote DNA
Transduction
• Some viruses can incorporate their genetic material into
the host’s chromosome
• When the virus is transferred from host to host, some of
the host’s genes can be transferred along with the virus’s
genes into the next host.
Transduction
Bacterium with
its chromosome
containing viral
(Prophage)
DNA
Prophage
DNA excised
from host
chromosome
Phage infects new
bacterium bringing
with it DNA from
the previous one
Phage DNA
inserts into new
host chromosome
along with
bacterial genes