Download The Search for the Genetic Material

Chapter 13
The Molecular Basis
of Inheritance
Question?
• Traits are inherited on
chromosomes, but what in the
chromosomes is the genetic
material?
• Two possibilities:
• Protein
• DNA
Qualifications
• Protein:
• very complex.
• high specificity of function.
• DNA:
• simple.
• not much known about it (early
1900’s).
For testing:
• Name(s) of experimenters
• Outline of the experiment
• Result of the experiment and
the importance of the result
Griffith - 1928
• Pneumonia in mice.
• Two strains:
• S - pathogenic
• R - harmless
Griffith’s Experiment
Result
• Something turned the R cells
into S cells.
• Transformation - the
assimilation of external genetic
material by a cell.
Problem
• Griffith used heat.
• Heat denatures proteins.
• So could proteins be the
genetic material?
• DNA - heat stable.
• Griffith’s results contrary to
accepted views.
Avery, McCarty and MacLeod
- 1944
• Repeated Griffith’s experiments,
but added specific fractions of S
cells.
• Result - only DNA transformed
R cells into S cells.
• Result - not believed.
Hershey & Chase -1952
• Genetic information of a virus or
phage.
• Phage - virus that attacks
bacteria and reprograms host to
produce more viruses.
Bacteria with Phages
Phage Components
• Two main chemicals:
• Protein
• DNA
• Which material is transferred to
the host?
Used Tracers
• Protein - CHONS, can trace
with 35S.
• DNA - CHONP, can trace with
32P.
Experiment
• Used phages labeled with one
tracer or the other and looked to
see which tracer entered the
bacteria cells.
Result
• DNA enters the host cell, but
the protein did not.
• Therefore:
DNA is the genetic material.
Picture Proof
Chargaff - 1947
• Studied the chemical
composition of DNA.
• Found that the nucleotides were
in certain ratios.
Chargaff’s Rule
• A=T
• G=C
• Example: in humans,
A = 30.9%
T = 29.4%
G = 19.9%
C = 19.8%
Why?
• Not known until Watson and
Crick worked out the structure
of DNA.
Watson and Crick - 1953
• Used X-ray crystallography data
(from Rosalind Franklin)
• Used model building.
• Result - Double Helix Model of
DNA structure.
• (One page paper, 1953).
Rosalind Franklin
Book & Movies
• “The Double Helix” by James
Watson- His account of the
discovery of the shape of DNA
• Movie – The Double Helix
DNA Composition
• Deoxyribose Sugar (5-C)
• Phosphate
• Nitrogen Bases:
• Purines
• Pyrimidines
DNA Backbone
• Polymer of sugar-phosphate.
• 2 backbones present.
Nitrogen Bases
• Bridge the backbones together.
• Purine + Pyrimidine = 3 rings.
• Constant distance between the
2 backbones.
• Held together by H-bonds.
Chargaff’s Rule
• Explained by double helix
model.
• A = T, 3 ring distance.
• G = C, 3 ring distance.
Watson and Crick
• Published a second paper
(1954) that speculated on the
way DNA replicates.
• Proof of replication given by
others.
Replication
• The process of making more
DNA from DNA.
• Problem: when cells replicate,
the genome must be copied
exactly.
• How is this done?
Models for DNA Replication
• Conservative - one old strand,
one new strand.
• Semiconservative - each strand
is 1/2 old, 1/2 new.
• Dispersive - strands are
mixtures of old and new.
Replication Models
Meselson – Stahl, late 1950’s
• Grew bacteria on two isotopes
of N.
• Started on 15N, switched to
14N.
• Looked at weight of DNA after
one, then 2 rounds of
replication.
Results
• Confirmed the
Semiconservative Model of DNA
replication.
Replication - Preview
• DNA splits by breaking the Hbonds between the backbones.
• Then DNA builds the missing
backbone using the bases on
the old backbone as a template.
Origins of Replication
• Specific sites on the DNA
molecule that starts replication.
• Recognized by a specific DNA
base sequence.
Prokaryotic
• Circular DNA.
• 1 origin site.
• Replication runs in both
directions from the origin site.
Eukaryotic Cells
• Many origin sites.
• Replication bubbles fuse to form
new DNA strands.
DNA Elongation
• By DNA Polymerases such as
DNA pol III
• Adds DNA triphosphate
monomers to the growing
replication strand.
• Matches A to T and G to C.
Energy for Replication
• From the triphosphate
monomers.
• Loses two phosphates as each
monomer is added.
Problem of Antiparallel DNA
• The two DNA strands run
antiparallel to each other.
• DNA can only elongate in the
5’--> 3’ direction.
Leading Strand
• Continuous replication toward
the replication fork in the 5’-->3’
direction.
Priming
• DNA pol III cannot initiate DNA
synthesis.
• Nucleotides can be added only
to an existing chain called a
Primer.
Primer
• Make of RNA.
• 10 nucleotides long.
• Added to DNA by an enzyme
called Primase.
• DNA is then added to the RNA
primer.
Priming
• A primer is needed for each
DNA elongation site.
Lagging Strand
• Discontinuous synthesis away
from the replication fork.
• Replicated in short segments as
more template becomes
opened up.
Okazaki Fragments
• Short segments (100-200
bases) that are made on the
lagging strand.
• All Okazaki fragments must be
primed.
• RNA primer is removed after
DNA is added.
Enzymes
• DNA pol I - replaces RNA
primers with DNA nucleotides.
• DNA Ligase - joins all DNA
fragments together.
Other Proteins in Replication
• Topoisomerase – relieves
strain ahead of replication
forks.
• Helicase - unwinds the DNA
double helix.
• Single-Strand Binding Proteins
- help hold the DNA strands
apart.
Video
• http://highered.mcgrawhill.com/olcweb/cgi/pluginpop.cgi?i
t=swf::535::535::/sites/dl/free/0072
437316/120076/micro04.swf::DNA
%20Replication%20Fork
Video
• http://www.bing.com/videos/sear
ch?q=YouTube+DNA+Replicatio
n+Process&Form=VQFRVP&adl
t=strict#view=detail&mid=1B8F7
75F1A027059094E1B8F775F1A
027059094E
DNA Replication Error Rate
• 1 in 1 billion base pairs.
• About 3 mistakes in our DNA
each time it’s replicated.
Reasons for Accuracy
• DNA pol III self-checks and
corrects mismatches.
• DNA Repair Enzymes - a family
of enzymes that checks and
corrects DNA.
DNA Repair
• Over 130 different DNA repair
enzymes known.
• Failure to repair may lead to
Cancer or other health
problems.
Example:
• Xeroderma Pigmentosum Genetic condition where a DNA
repair enzyme doesn’t work.
• UV light causes damage, which
can lead to cancer.
Xeroderma Pigmentosum
Cancer
Protected from UV
Thymine Dimers
• T-T binding from side to side
causing a bubble in DNA
backbone.
• Often caused by UV light.
Excision Repair
• Cuts out the damaged DNA.
• DNA Polymerase fills in the
excised area with new bases.
• DNA Ligase seals the
backbone.
Problem - ends of DNA
• DNA Polymerase can only add
nucleotides in the 5’--->3’
direction.
• It can’t complete the ends of the
DNA strand.
Result
• DNA gets shorter and shorter
with each round of replication.
Telomeres
• Repeating units of TTAGGG
(100- 1000 X) at the end of the
DNA strand (chromosome)
• Protects DNA from unwinding
and sticking together.
• Telomeres shorten with each
DNA replication.
Telomeres
Telomeres
• Serve as a “clock” to count how
many times DNA has replicated.
• When the telomeres are too short,
the cell dies by apoptosis.
Implication
• Telomeres are involved with the
aging process.
• Limits how many times a cell line
can divide.
Telomerase
• Enzyme that uses RNA to
rebuild telomeres.
• Can make cells “immortal”.
• Found in cancer cells.
• Found in germ cells.
• Limited activity in active cells
such as skin cells
Comment
• Control of Telomerase may stop
cancer, or extend the life span.
NEWS FLASH
• The DNA of Telomers is
actually used to build proteins.
• These proteins seem to impede
telomerase.
• Feedback Loop??
Chromatin Packing
1. Nucleosomes
2. 30-nm Chromatin Fibers
3. Looped Domains
4. Chromosomes
Focus on #1 & 4
Nucleosomes
•
•
•
•
"Beads on a String”.
DNA wound on a protein core.
Packaging for DNA.
Controls gene reading
Protein Core
• Two molecules of four types of
Histone proteins.
• H1- 5th type of Histone protein
attaches the DNA to the outside
of the core.
Chromosomes
• Large units of DNA.
• Similar to "Chapters" in the
Book of Life.
Summary
• Know the Scientists and their
experiments.
• Why DNA is an excellent
genetic material.
• How DNA replicates.
• Problems in replication.
• Chromatin packing