Download Ch7 microbgeneticspart1HOLrg

10/21/13
Genetics: Chapter 7
What is genetics?
•  The science of heredity; includes the study
of genes, how they carry information, how
they are replicated, how they are expressed
1
10/21/13
Prokaryotic DNA replication,
trascription and translation
Gene Expression
DNA
DNA Replication
Duplicates the DNA
moleculeso its encoded
information can be passed
on to the next generation.
Transcription
Copies the information in
DNA into RNA.
RNA
Translation
Interprets the information
carried by RNA to synthesize
the encoded protein.
Protein
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
DNA structure: single subunit is a
deoxyribonucleic acid—3 parts
Adenosine
NH2
N
O
O
O
O
–
P
O
P
O
P
O
–
O
–
High-energy
bonds
O
–
N
O
CH2
N
O
N
Adenine
OH
OH
Ribose
Where else have you seen this molecule?
2
10/21/13
RNA vs.DNACH OH
O
OH
C
O
C
H
H
C C
OH
H
C
C H
C
OH
OH OH
Ribose
C
OH
C
OH
H
H
CH OH
O
O
H
C
O
C
H
H
C C
H
H
C
C
H
C
OH
OH H
Deoxyribose
C
OH
C
OH
H
H
5
2
1
H
H
H
H
1
4
2
3
2
3
4
5
5
2
1
H
H
H
H
4
2
1
3
3
4
5
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Purines
(double ring)
Pyrimidines
(single ring)
N
N
H
CH3
N
H
N
H
Adenine (A)
(both DNA and RNA)
O
N
N
H
N
H
N
O
O
NH2
H
N
O
H
Thymine (T)
(DNA only)
H
H
N
N
H
O
H
Uracil (U)
(RNA only)
NH2
H
H
NH2
H
Guanine (G)
(both DNA and RNA)
N
H
N
N
O
H
Cytosine (C)
(both DNA and RNA)
3
10/21/13
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Sugarphosphate
backbone
Nucleobases
G
P
5' phosphate
1
4
3
2
A
P
5
3
P
1
4
Nucleotide
5
2
C
5
1
4
3
2
T
P
5
The 3' carbon of
one nucleotide is
linked to the 5' carbon
of the next nucleotide
via a phosphate group.
1
4
3
OH
2
3' hydroxyl
Double stranded DNA forms a
double helix
4
10/21/13
DNA double strand
5′ phosphate
5′ end
3′ hydroxyl
3′ end
HO
T
C
G
A
G
Sugar
C
Nucleotide
O
O
Sugar
O
P
P
T
Sugar
O
Sugar
P
O
P
C
Sugar
Sugar
O
P
DNA
O
P
G
Sugar
O
Sugar
P
O
Sugar
P
A
Sugar
P
Base
pairs
O
P
Notice the strands run
in the opposite
direction relative to
eachother-
Hydrogen
bonds
HO
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
3′ hydroxyl
3′ end
5′ end
5′ phosphate
5
10/21/13
DNA is synthesized in one
direction. New units added to
the 3’ end only. So, direction
is from the 5’ to 3’ BECAUSE, hydroxyl group
is on the 3’ carbon of the
pentose ring.
DNA Replication 6
10/21/13
Enzymes necessary for DNA
replication
• 
• 
• 
• 
• 
Primase
DNA Polymerase
DNA gyrase
DNA ligase
Helicase
DNA Polymerase adds nucleotides to the 3’ end
Template strand
5′
DNA Replication
3′
T A C G G T A C T A G T A G T A G T C G A T T C G A A
T C A T C A T C A G C T A A G C T T
Direction
of synthesis
3′
A
5′
DNA polymerase
P
P
O
A
P
O
P
P
A
G
T
C
O
O
OH
P
O
T
O
OH
P
New strand
P
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
7
10/21/13
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Replication forks
3′
5′
1
A helicase unzips
the two strands of DNA.
Helicase
Leading
strand
5′
3′
5
RNA primer
DNA polymerase adds
nucleotides onto the 3′
end of the strand.
5′
3′
5′
2
Synthesis of the leading strand
proceeds continuously as fresh
template is exposed.
5′
5′
3′
Okazaki fragment
of the lagging strand
Primase synthesizes
the RNA primer.
3
Synthesis of the lagging strand must be reinitiated as more
template is exposed. Each time synthesis is reinitiated,
a new RNA primer must be made. Discontinuous synthesis
generates Okazaki fragments.
5′
3′
5′
4
As DNA polymerase adds nucleotides
to the 3′ end of one Okazaki fragment,
it encounters the 5′ end of another.
A different type of DNA polymerase
then removes the RNA primer
nucleotides and simultaneously
replaces them with deoxynucleotides.
3′
5′
5
DNA ligase seals
the gaps between
Okazaki fragments
by forming a covalent
bond between them.
DNA ligase
3′
5′
8
10/21/13
DNA
replication:
detail
DNA replication…closer look
9
10/21/13
Gene Expression…why is it
important?
•  Transcription
•  Translation
Transcription: DNA to RNA
•  Requires an enzyme…..
•  RNA nucleotides
•  Base pairing rules for building RNA from a
DNA template
•  Process proceeds in the direction 5 --->3 •  Process begins at the promoter region and
ends at the terminator sequence
10
10/21/13
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Gene Expression
Transcription
Translation
Transcription: RNA is
transcribed from DNA
3′
Plus (+)
5′
strand
GCTGATGAT CCGCGTAGGTGC T
of DNA
CGAC T AC T AGGCGC AT C C ACGA
5′
Minus (–)
3′
strand
of DNA
3′
RNA
5′
GCUGAUGAUCC GCGUAGGUGCU
Transcription: Promoter orients direction
of transcription
11
10/21/13
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Gene Expression
Transcription
Transcription: RNA synthesis
Translation
5′
3′
3′
5′
Promoter
Terminator
RNA polymerase
5′
3′
3′
5′
1
Template
strand
Sigma
5′
2
3′
3′
5′
Promoter
Initiation
RNA polymerase binds to the
promoter and melts a short
stretch of DNA.
5′
RNA
Elongation
Sigma factor dissociates from RNA
polymerase, leaving the core
enzyme to complete transcription.
RNA is synthesized in the 5′ to 3′
direction as the enzyme adds
nucleotides to the 3′ end of
the growing chain.
A CUG
G
C T G AC
3
5′
3′
3′
5′
Promoter
Termination
When RNA polymerase
encounters a terminator, it falls
off the template and releases
the newly synthesized RNA.
5′
RNA polymerase
dissociates from template.
What are the possible products
from transcription?
•  Messenger RNA (mRNA)
•  Transfer RNA (tRNA)
•  Ribosomal RNA (rRNA)
12
10/21/13
Fig. 7.3
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Protein-encoding gene
DNA
rRNA gene
tRNA gene
Transcription
Messenger RNA (mRNA) Ribosomal RNA (rRNA) Transfer RNA (tRNA)
Translation
Protein
Quick check…..
•  Do we have a protein yet?
•  What have we made? •  What is next?
13
10/21/13
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Gene Expression
Translation:RNA to
Protein
Transcription
Translation
Region translated
5′
3′
mRNA
Ribosome- Start
binding site codon
Stop
codon
Translation
Protein
Phe
Ser His
Cys
Tyr
Ser
Pro
Ala
Gln
Ser Met
Tyr
Gly
Glu Val
Leu
Amino acid
P ro
•  What is needed for the process?
–  mRNA
–  Ribosomes
–  Amino acids
–  tRNA
Hydrogen bond
tRNA
Anticodon
G G C
C C G
5′
mRNA
3′
(a)
Codon
P ro
tRNA
G G C
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
(b)
Anticodon
14
10/21/13
Translation: reading frame
determines the protein
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Non-polar
Amino acids
H3
C
CH3
H3
C
CH3
CH2
CH3
C
H
H3
N+
C
COO –
H3
N+
C
COO–
H3
N+
C
COO–
H
H
Glycine (Gly; G)
Alanine (Ala; A)
CH3
CH
H
H3
N+
H
Valine (Val; V)
C
COO–
CH2
C
H
H2
C
CH2
H3
N+
C
COO–
H2
N+
C
COO–
H
H
H
Leucine (Leu; L)
Isoleucine (IIe; I)
Proline (Pro; P)
CH3
H
N
S
SH
CH2
H 3N +
CH2
C
COO–
H 3N +
H
Phenylalanine (Phe; F)
CH2
H3C
CH2
CH2
C
COO–
H 3N +
CH2
C
COO–
H 3N +
C
COO –
H
H
H
Tryptophan (Trp; W)
Cysteine (Cys; C)
Methionine (Met; M)
Polar/hydrophilic (uncharged)
O
O
OH
CH2
H 3N +
C
COO–
C
HO CH3
CH
H 3N +
H
H 3N +
OH
NH2
C
CH2
CH2
CH2
C
COO–
H
Serine (Ser; S)
N
H
2
C
COO–
H 3N +
H
C
COO–
CH2
H 3N +
H
Threonine (Thr; T) Asparagine (Asn; N) Glutamine (Gln; Q)
C
COO–
H
Tyrosine (Tyr; Y)
Polar/hydrophilic (charged)
Basic
NH2
C
NH2+
Acidic
NH3+
O
–
O
O
–
O
C
CH2
H 3N +
C
COO–
H
Aspartic acid (Asp; D)
HN
CH2
C
CH2
CH2
H 3N +
C
COO –
H
Glutamic acid (Glu; E)
H 3N +
C
COO–
H
Histidine (His; H)
CH2
NH
CH2
CH2
CH2
CH2
CH2
CH2
H 3N +
C
COO –
H
Lysine (Lys; K)
H 3N +
C
COO –
H
Arginine (Arg; R)
15
10/21/13
Fig. 2.13
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
R
Side chain—
R is the general
designation for a side chain
H
Amino group—
positively charged
at neutral pH
O
H
N+
C
C
H
H
O–
Carboxyl group—
negatively charged
at neutral pH
The Genetic code
16
10/21/13
Structure of Ribosomes
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
f-Met
P-site
E-site
Translation
Initiation
The initiating tRNA, carrying the amino
acid f-Met, base-pairs with the start
codon and occupies the P-site.
A-site
U A C
A U G C C G U A C G A A G A U U A C U A G G A U
3′
5′
mRNA
f-Met
Pro
A tRNA that recognizes the next codon
then fills the unoccupied A-site.
U A C G G C
A U G C C G U A C G A A G A U U A C U A G G A U
3′
5′
Peptide bond
f-Met
Pro
The ribosome catalyzes the joining of the
amino acid carried by the tRNA in the
P-site to the one carried by the tRNA in
the A-site.
5′
U A C G G C
A U G C C G U A C G A A G A U U A C U A G G A U
3′
(a)
17
10/21/13
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
R
R
H
H
H
O
N+
C
H
H
+
C
H
O–
Amino acid
N+
C
H
H
C
O–
Amino acid
Dehydration
synthesis
H2O
R
R
O
H
H
O
N+
C
H
H
C
O
N
C
H
H
C
O–
18
10/21/13
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Ty
r
f-Met
Pro
U
A
C
P-site
E-site
A-site
A
U
G
G G C
A U G C C G U A C G A A G A U U A C U A G G A U
5′
3′
Ribosome moves along mRNA.
Elongation
The ribosome advances a distance of
one codon. The tRNA that occupied the
P-site exits through the E-site and the
tRNA that was in the A-site occupies the
P-site. A tRNA that recognizes the next
codon quickly fills the empty A-site.
f-Met
Gl
u
Pro
Tyr
The ribosome continues advancing
down the mRNA in the 5′ to 3′ direction,
moving one codon at a time.
C
G G
C
U
U
A U G
A U G C C G U A C G A A G A U U A C U A G G A U
5′
3′
(b)
f-Met
Pro
Tyr
Glu
Asp
Tyr
C
U
A
A U G
A U G C C G U A C G A A G A U U A C U A G G A U
5′
3′
Termination
Translation continues until a stop
codon is reached, signaling the end
of the process. No tRNA molecules
recognize a stop codon.
The components dissemble, releasing
the newly formed polypeptide.
Pro
f-Met
Tyr
Glu
Asp
Tyr
(c)
Both processes occur at the same
time in bacteria…why??
19
10/21/13
Eukaryotic cells differ in
transcription and translation
•  Ribosomes are different size
•  5 end of mRNA has cap (methylated
guanine)
•  3 end of mRNA has poly A tail
•  Introns are excised, exons spliced together
•  Translation is monocystronic
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Eukaryotic transcription
Exon
Intron
Exon
Eukaryotic DNA contains
introns, which interrupt
coding regions (exons).
Intron
Exon
Eukaryotic DNA
Transcription generates
pre-mRNA (precursor mRNA)
that contains introns. A cap
and poly A tail are then added.
Poly A tail
Cap
Pre-mRNA
Splicing removes introns to
create functional mRNA.
mRNA
mRNA is transported out of
the nucleus to be translated
In the cytoplasm.
20
10/21/13
Is it important to regulate protein
synthesis?
•  Yes!
•  Three types of protein regulation
–  Enyme inhibition (feedback inhibition)
–  Repression (tryptophan operon)
–  Induction (lactose operon)
Are all genes under regulation?
•  No!
•  Genes to produce enzymes for glucose
metabolism are constitutive (always made)
•  Other genes are induced…only made when
needed
•  Other genes are repressed…turned off
when not needed
21
10/21/13
Models for transcriptional
regulation with repressors
Transcriptional regulation by
activators
22
10/21/13
Lactose Operon as a model
•  Used to understand control of gene
expression in bacteria
•  Operon consists of three genes needed to
degrade lactose
•  Repressor gene(codes for repressor protein)
outside of operon coding region inhibits
transcription unless something else bind to
the repressor protein
Lactose Operon
23
10/21/13
Diauxic growth curve of E. coli
24
10/21/13
What conditions are needed for the lactose
operon to be turned on ?
• 
• 
• 
• 
No glucose
Lactose present
Increasing levels of cAMP
cAMP binds to CAP, then complex binds
next to lactose operon promoter at the
activator region
•  RNA polymerase binds to promoter
How do organisms adapt to other
changes in their environment?
•  Some organisms turn genes on/off as
needed
•  Some organisms alter gene expression
25
10/21/13
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Gene regulation
systems in bacteria
•  Signal transduction
–  Two component
regulatory system
Environmental stimulus
Sensor
protein
Response
regulator
The sensor protein spans the cytoplasmic membrane.
The response regulator is a protein inside the cell.
P
P
In response to a specific change in the environment, the
sensor phosphorylates a region on its internal portion.
The phosphate group is transferred to the response
regulator, which can then turn genes on or off, depending
on the system.
•  Signal transduction
- Quorum sensing
Bacterial cell
When few cells are present, the
concentration of the signaling
molecule is low.
Signaling
molecule
When many cells are present, the
signaling molecule reaches a
concentration high enough to induce
the expression of certain genes.
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
26
10/21/13
Gene expression is influenced by
natural selection
•  Random changes enhance survival of some
cells in population
•  Antigenic variation of pathogens
•  Phase variation
–  Switching on/off of certain genes
27