Chapter 11 DNA: The Carrier of Genetic Information
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Transcript of Chapter 11 DNA: The Carrier of Genetic Information
Chapter 11DNA: The Carrier of Genetic
Information
Experiments in DNA
• ???Protein as the genetic material• 20 AA – many different combinations = unique
properties• Genes control protein synthesis• DNA and RNA – only 4 nucleotides = dull
Experiments in DNA• Frederick Griffith – 1928
– Bacteria – pneumococcus – 2 strains– (S) smooth strain – virulent (lethal)
• Mice – pneumonia - death
– (R) rough strain – avirulent• Mice survive
– Heat killed (S) strain• Mice survive
– Heat killed (S) + live (R)• Mice died• Found living (S) in dead mice
• Griffith continued– transformation - type of permanent genetic
change where the properties of 1 strain of dead cells are conferred on a different strain of living cells
– “transforming principle” was transferred from dead to living cells
Fig. 16-2
Living S cells (control)
Living R cells (control)
Heat-killed S cells (control)
Mixture of heat-killed S cells and living R cells
Mouse diesMouse dies Mouse healthy Mouse healthy
Living S cells
RESULTS
EXPERIMENT
• Avery, MacLeod, McCarty - 1944– Identified Griffith’s transforming principle as DNA– Live (R) + purified DNA from (S) R cells
transformed– R + (S) DNA die– R = (S) protein live
– DNA responsible for transformation– Really?
• Hershey and Chase – 1952– Bacteriophages– Radioactive labels
• Viral protein – sulfur• Viral DNA - phosphorus
– infect bacteria, agitate in blender, centrifuge– Found
• Sulfur sample – all radioactivity in supernatant (not cells)• Phosphorus sample – radioactivity in pellet (inside cells)
– SO – bacteriophages inject DNA into bacteria, leaving protein on outside
– DNA = hereditary material
Fig. 16-3
Bacterial cell
Phage head
Tail sheath
Tail fiber
DNA
100
nm
Fig. 16-4-3
EXPERIMENT
Phage
DNA
Bacterial cell
Radioactive protein
Radioactive DNA
Batch 1: radioactive sulfur (35S)
Batch 2: radioactive phosphorus (32P)
Empty protein shell
Phage DNA
Centrifuge
Centrifuge
Pellet
Pellet (bacterial cells and contents)
Radioactivity (phage protein) in liquid
Radioactivity (phage DNA) in pellet
• Rosalind Franklin (in lab of Wilkins)– X-ray diffraction on crystals of purified DNA– (X-ray crystallography)– Determine distance between atoms of molecules
arranged in a regular, repeating crystalline structure
• Helix structure• Nucleotide bases like rungs on ladder
Fig. 16-6
(a) Rosalind Franklin (b) Franklin’s X-ray diffraction photograph of DNA
• James Watson and Francis Crick – 1953– Model for DNA structure = double helix– DNA now widely accepted as genetic material– Took all available info on DNA and put together– Showed –
• DNA can carry info for proteins• Serve as own template for replication
Structure of DNA
• Nucleotides– Deoxyribose– Phosphate– Nitrogenous base (ATCG)
• Purines – adenine, guanine – 2 rings• Pyrimidines – thymine, cytosine – 1 ring
– covalent bonds link = sugar-phosphate backbone• 3’ C of sugar bonded to 5’ phosphate = phophodiester
linkage• 5’ end – 5’ C attached to phosphate• 3’ end – 3’ C attached to hydroxyl
Chargaff - 1950
• # purines = # pyrimidines– #A = #T– #C = # G
• Each cross rung of ladder– 1 purine + 1 pyrimidine
Fig. 16-UN1
Purine + purine: too wide
Pyrimidine + pyrimidine: too narrow
Purine + pyrimidine: width consistent with X-ray data
• Hydrogen bonding between N bases• A-T = 2 H bonds• G-C = 3 H bonds• Complementary base pairs
• # possible sequences virtually unlimited• many genes, much info
Fig. 16-8
Cytosine (C)
Adenine (A) Thymine (T)
Guanine (G)
Fig. 16-5 Sugar–phosphate backbone
5 end
Nitrogenous
bases
Thymine (T)
Adenine (A)
Cytosine (C)
Guanine (G)
DNA nucleotide
Sugar (deoxyribose)
3 end
Phosphate
Fig. 16-7a
Hydrogen bond 3 end
5 end
3.4 nm
0.34 nm3 end
5 end
(b) Partial chemical structure(a) Key features of DNA structure
1 nm
DNA Replication
• Semiconservative – each strand of DNA is template to make opposite new strand
• Meselson and Stahl– E. coli and isotopes of N– 15N – heavy/dense; 14N “normal”– Bacteria with 15N in DNA replicated with medium
having 14N– Centrifuge – Supports semiconservative model
• Explains how mutagens can be passed on
Fig. 16-11a
EXPERIMENT
RESULTS
1
3
2
4
Bacteria cultured in medium containing 15N
Bacteria transferred to medium containing 14N
DNA sample centrifuged after 20 min (after first application)
DNA sample centrifuged after 20 min (after second replication)
Less dense
More dense
Fig. 16-9-3
A T
GC
T A
TA
G C
(a) Parent molecule
A T
GC
T A
TAG C
(c) “Daughter” DNA molecules, each consisting of one parental strand and one new strand
(b) Separation of strands
A T
GC
T A
TA
G C
A T
GC
T A
TAG C
Fig. 16-10Parent cell
First replication
Second replication
(a) Conservative model
(b) Semiconserva- tive model
(c) Dispersive model
Steps of DNA Replication
• 1. DNA helicase – • 2. Helix-destabilizing proteins – • 3. Topoisomerases – • 4. RNA primer – • 5. DNA polymerase – • 6. Origin of replication –
– Leading strand– Lagging strand
• 7. DNA Ligase
Leading Strand
Fig. 16-12b
0.25 µm
Origin of replication Double-stranded DNA molecule
Parental (template) strandDaughter (new) strand
Bubble Replication fork
Two daughter DNA molecules
(b) Origins of replication in eukaryotes
Fig. 16-14
A
C
T
G
G
G
GC
C C
C
C
A
A
AT
T
T
New strand 5 end
Template strand 3 end 5 end 3 end
3 end
5 end5 end
3 end
BaseSugar
Phosphate
Nucleoside triphosphate
Pyrophosphate
DNA polymerase
Fig. 16-15a
Overview
Leading strand
Leading strandLagging strand
Lagging strandOrigin of replication
Primer
Overall directions of replication
Fig. 16-17
OverviewOrigin of replicationLeading strand
Leading strand
Lagging strand
Lagging strandOverall
directions of
replicationLeading strand
Lagging strand
Helicase
Parental DNA
DNA pol IIIPrimerPrimase
DNA ligase
DNA pol IIIDNA pol I
Single-strand
binding protein
53
5
55
5
3
3
3313 2
4
Telomeres
• Telomeres – caps end of chromosome; short non-coding sequences repeated many times
• Cell can divide many times before losing crucial info
• Lagging strand is discontinuous, so DNA polymerase unable to complete replication , leaving small part unreplicated small part lost with each cycle
Fig. 16-19Ends of parental DNA strands
Leading strandLagging strand
Lagging strand
Last fragment Previous fragment
Parental strand
RNA primer
Removal of primers and replacement with DNA where a 3 end is available
Second round of replication
New leading strand
New lagging strand
Further rounds of replication
Shorter and shorter daughter molecules
5
3
3
3
3
3
5
5
5
5