In This Lesson:
Unit 6
DNA Structure
and Replication
(Lesson 1 of 3)
Today is Wednesday,
December 23rd, 2015
Pre-Class:
Name as many enzymes as you can that are involved in
DNA replication.
Get a small paper towel, too.
Today’s Agenda
•
•
•
•
Midterm Analysis.
DNA history.
DNA structure.
DNA replication.
– Also known as a look into the details of S phase.
• A DNA pickup line.
• Where is this in my book?
– Chapter 16.
By the end of this lesson…
• You should be able to describe the structure
of DNA in detail.
• You should be able to narrate the replication
of a DNA molecule including all enzymes used
therein.
• You should be able to describe the cell’s
mechanism for detecting and repairing
mutations.
Midterm Analysis
• Now that everyone’s done with the midterm,
you’re each going to review your own work to
see what went right and what may have gone
wrong.
• On the accompanying “worksheet,” answer
the questions honestly and completely.
• You’ll turn it in today but will get it back when
the AP Exam and final are on the horizon.
Let’s Begin at the Beginning
• Challenge Questions!
• DNA Base Pairs worksheet.
DNA Worksheet
• Problems 1 and 2 on your DNA worksheet:
1:
1:
AC
TG
2: G A A G G C G T T
DNA Worksheet
• Problems 3, 4, and 5 on your worksheet:
3: T T G C A A G T C
4:
5. Always three rings (keeps the same width).
DNA Worksheet
• Problems 6 and 7 on your worksheet:
6: Sugar/Phosphate Backbone
7: A ↔ T, C ↔ G
7: Purines – 2 rings, Pyrimidines – 1 ring
The Historical Perspective
• Who discovered DNA?
– You’re probably thinking…Watson and Crick?
• That’s accurate…but only…kind of.
• The elucidation of DNA and its structure is a
fantastic study of science at work, and the way
“standing on the shoulders of giants” that
have come before lets us as humanity leap
forward into the bold future of biology.
– Preach!
The Historical Perspective
• 1869: Friedrich Miescher
– Miescher discovered DNA
while analyzing pus from
discarded bandages.
– He named the substance
“nuclein” but did not realize he
was looking at the origin of
evolution.
• Must have been like when you
realize you saw a celebrity after
he’s gone.
http://upload.wikimedia.org/wikipedia/commons/b/bc/Friedrich_Miescher.jpg
Johann Friedrich Miescher
The Historical Perspective
• 1908: Thomas Hunt Morgan
– Remember him?
– Morgan worked with Drosophila
and found that genes were
located on chromosomes.
– However, Morgan did not know
whether it was the DNA or the
histone proteins that are the
actual genes.
T.H. Morgan
The Historical Perspective
• 1928: Frederick Griffith
– Griffith worked with pneumonia-causing
Steptococcus bacteria in an attempt to find a
cure.
– He mixed harmless live bacteria with harmful
dead (killed by heat) bacteria.
– The once-harmless bacteria somehow took up
the harmful “transforming factor” of the
harmful bacteria.
• Something could be passed on, but it was unknown
what that “transforming factor” was.
– Mice injected with the harmless live bacteria
(after mixing) were killed.
Frederick Griffith
(in the hat)
The Historical Perspective
Griffith
• Griffith’s experiment:
Live,
Pathogenic
Bacteria
Live, NonPathogenic
Bacteria
Killed
Pathogenic
Bacteria
Mix of Killed
Pathogenic and
Live NonPathogenic
Bacteria
The Historical Perspective
• 1944: Oswald Avery, Maclyn McCarty, Colin MacLeod
– Refined the results of Griffith’s work by purifying protein and
DNA from Streptococcus and running the same experiment.
– Results? Proteins had no effect on mice, but DNA did.
Oswald Avery
Maclyn McCarty
Colin MacLeod
http://profiles.nlm.nih.gov/ps/access/CCAACA.jpg
http://media-1.web.britannica.com/eb-media/12/160212-004-5DF19B15.jpg http://www.nlm.nih.gov/visibleproofs/media/detailed/vi_a_204.jpg
The Historical Perspective
• 1947: Erwin Chargaff
– Discovered what is now known as
Chargaff’s Rules. In DNA:
• % of Adenine ≈ % of Thymine
• % of Cytosine ≈ % of Guanine
Relative Proportions (%) of Bases in DNA
Organism
A
T
G
C
Human
30.9 29.4 19.9
19.8
Chicken
28.8 29.2 20.5
21.5
Grasshopper 29.3 29.3 20.5
20.7
Sea Urchin
32.8 32.1 17.7
17.3
Wheat
27.3 27.1 22.7
22.8
Yeast
31.3 32.9 18.7
17.1
E. coli
24.7 23.6 26.0
25.7
Erwin Chargaff
The Historical Perspective
• 1952: Alfred Hershey and
Martha Chase
– The classic “blender
experiment.”
– The two used a type of virus that
infects bacteria (bacteriophage)
and created two groups:
– One labeled with 35S in protein.
– One labeled with 32P in DNA.
• Each of those isotopes is
radioactive and thus detectable.
Chase & Hershey
The Historical Perspective
Hershey-Chase
Bacteriophages are
grown in radioactive
cultures and tagged.
32P 
 35S
[protein]
[DNA]
Viruses infect
cells by
injecting DNA.
35S
(protein)
is found
outside the
cells.
Viruses infect
cells by
injecting DNA.
32P
(DNA) is
found inside
the cells.
The Historical Perspective
Hershey-Chase
• The Hershey-Chase experiment thus answers the
question of what was the “transforming factor”
from Griffith’s work.
– Since marked protein never made it into the cell, it
couldn’t be the protein in the chromatin.
– Since DNA did make it into the cell, that must have
been what allowed the harmless bacteria to transform.
• So where’d the blender come in?
– The blender was used to agitate the virions (virus
particles) off the cells to allow for study of just the
cells’ contents and not the tagged viruses.
The Historical Perspective
Hershey-Chase
The Historical Perspective
• 1953: Watson and Crick (and Franklin
and Wilkins)
Maurice Wilkins
Rosalind Franklin
Francis Crick
James Watson
– Rosalind Franklin and Maurice Wilkins
used X-ray crystallography to learn about
the structure of DNA.
– The image to the right shows a uniform Xshape, suggesting a helix shape with a
consistent width. So where do Watson
and Crick come in?
The Historical Perspective
• Watson and Crick used
the work of Franklin and
Wilkins to create models
of DNA, eventually
figuring out its structure.
– They still deserve credit,
but Wilkins and Franklin
deserve just as much.
Da Structurez
• So what comes out of all that work?
• The classic DNA structure: a double helix.
– Meaning it looks like that spiral staircase in Gattaca.
Coincidence?
• I think not.
http://media-cache-ec0.pinimg.com/736x/1a/02/bc/1a02bc5d5428bbdc64e793f61bf2b809.jpg
DNA Structure Review
• DNA is a nucleic acid, a long
string of nucleotides.
• DNA takes the shape of a
double-helix.
• There are four kinds of
nucleotides:
– Adenine
– Cytosine
– Guanine
– Thymine
http://ghr.nlm.nih.gov/handbook/illustrations/dnastructure.jpg
Nucleotide Structure Review
• Each nucleotide has a:
– Sugar molecule with 5-carbons (pentose)
• Deoxyribose in DNA
• Ribose in RNA
– Phosphate group
• Phosphorous-based molecule
– Nitrogenous base (makes the nucleotide unique)
•
•
•
•
Adenine
Thymine
Cytosine
Guanine
Nucleotide Structure Review
Guanine
Adenine
Thymine
Cytosine
http://www.biologyjunction.com/images/nucleotide1.jpg
Nucleotide Structure Review
• More “scientific”
Nucleotides and Nucleosides
• Just so you know, you’ll occasionally hear of a
nucleoside.
• The only difference between a nucleoside and
a nucleotide is that a nucleoside is just a sugar
and nitrogenous base – no phosphate group.
DNA Structure Review
• Surrounding the base pairs and
forming the sides of the
“ladder” is a sugar-phosphate
backbone.
• The backbone is made of a sugar
(deoxyribose) and a phosphate
group, alternating and in reverse
order from the other strand.
– Backbone is linked by
phosphodiester bonds.
– The end of DNA with the
phosphate on top is the 5’ (“five
prime”) end.
– The other end of the backbone is
the 3’ (“three prime”) end.
http://ghr.nlm.nih.gov/handbook/illustrations/dnastructure.jpg
3’ and 5’? Huh?
• 3’ and 5’ get their names from
the pentose sugar’s carbon
atoms.
• Each carbon in pentose is
numbered and has a specific job
in the formation of DNA.
– Carbon 1 = base attachment
– Carbon 2 = oxygen (ribose) or
not (deoxyribose)?
– Carbon 3 = another nucleotide
attachment
– Carbon 4 = completes ring
– Carbon 5 = phosphate
attachment
• This is important.
http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/P/Pentose.gif
http://www.synapses.co.uk/genetics/pentose1.gif
DNA Structure Review
• DNA stands for
Deoxyribonucleic Acid.
• By hydrogen bonds,
cytosine bonds to
guanine and adenine to
thymine.
• A↔T
• C↔G
http://ghr.nlm.nih.gov/handbook/illustrations/dnastructure.jpg
One more time, because “important.”
 Oxygen, not
a zero.
5’
3’
DNA Unwound
P
P
---H---
Thymine
PD Bond
Adenine
Deoxyribose
Deoxyribose
PD Bond
---H---
Guanine
Cytosine
Deoxyribose
Deoxyribose
PD Bond
P
P
PD Bond
---H---
Adenine
Thymine
Deoxyribose
Deoxyribose
5’
P
3’
Antiparallel Strands
• As seen in the image to the right,
the two strands of DNA run
antiparallel to one another.
– One is “upside down.”
• At the 5’ end of each DNA strand
there is a phosphate group.
• At the 3’ end of each DNA strand
there is a hydroxyl (-OH) group.
Bonding in DNA
Phosphodiester Bond
• The two complementary
strands of DNA are linked
by hydrogen bonds.
– Base to base.
• Each nucleotide in a
sugar-phosphate
backbone is linked by a
phosphodiester bond.
– Phosphate group to 3’ C.
Hydrogen Bond
Linking Nucleotides
• Phosphodiester
bonds, linking
nucleotides, are
formed…how?
– By dehydration
synthesis, of
course!
– More on this later.
Purines and Pyrimidines
• Adenine and guanine are
purines and have a doublering structure.
• Cytosine and thymine are
pyrimidines and have a singlering structure.
– A purine always bonds to a
pyrimidine.
– This ensures that the width of
the double helix is constant.
• How can we remember this
one?
DNA Replication
• There comes a time in (almost) every DNA
molecule’s life when it needs to be replicated
(copied).
– That time would be S phase.
• Here’s the general process:
– Unwind the double-helix.
– Break the hydrogen bonds (“unzip” the DNA).
– Use enzymes to replace base pairs on each side.
Thymine
Deoxyribose
Adenine
Thymine
Replication
P
P
P apart
PDNA Polymerase makes
DNA move
DNAnew
Helicase
breaks
H-bonds
Strands
---H---
Thymine
P
Deoxyribose
Deoxyribose
P
Deoxyribose
P
Thymine
H
P
Guanine
Adenine
DeoxyH
ribose
Deoxyribose
Deoxyribose
P
Cytosine
Adenine
---H---
Guanine
Cytosine
Deoxyribose
DeoxyH
ribose
H
H
P
---H---
Deoxyribose
Adenine
Deoxyribose
DeoxyH
ribose
Looks like this… [IMPORTANT]
Note that even
though there
are two
strands
forming down
here, each is
only “half”
new.
The “old”
strand is
sometimes
known as the
template
strand
because it’s a
model for the
new one.
Looks like this… [IMPORTANT]
http://online.santarosa.edu/homepage/cgalt/BIO10-Stuff/Ch10-Protein_Synthesis/DNA-Replication-Animation.gif
KindaPOGIL
• We’re going to do a segment of a POGIL now.
• DNA Structure and Replication [Model 2 only]
The Historical Perspective
• One more experiment, but it’s a good one.
– It’s actually known as “The Most Beautiful Experiment in
Biology.”
• In 1958, Meselson & Stahl set out to determine how
DNA is copied.
– Conservative
• “Parent” DNA exists in whole after copying.
– Semi-conservative
• “Parent” DNA is divided into two whole strands – one in each of
the daughter molecules.
– Dispersive
• “Parent” DNA is fragmented among the two daughter molecules.
• Which one do we now know to be correct?
DNA Replication Models
• Compare the models:
The Historical Perspective
• So how did Meselson and Stahl pull
this one off?
– They started by “labeling” nucleotides
in the parent bacterial DNA with 15N
(“heavy nitrogen”).
– New nucleotides were labeled with 14N.
Franklin Stahl
Matthew Meselson
http://www.g2conline.info/content/c16/16448/16448_stahloffice.jpg
http://library.cshl.edu/static/oh4/images_still/MatthewMeselson.png
The Historical Perspective
• After one replication, they found that the density of
the nucleotides was between that of 15N and 14N.
– So it can’t be conservative – that would have two
separate densities (15N and 14N).
The Historical Perspective
• After two replications, they found that the density of
the nucleotides was between that of 15N and 14N OR
equal to that of 14N.
– So it can’t be dispersive – that would have one density
slightly higher than the first replication.
Back to Replication…
• So how would you guess DNA replication is
actually achieved?
– As in, we’re talking about molecules here. How is
the molecular “work” done?
• Yep, enzymes. The repeat offenders of
“getting stuff done.”
• Can you remember any enzymes involved in
replication?
Replication Enzymes
•
•
•
•
•
Helicase
DNA Polymerase III (abbreviated pol III)
DNA Polymerase I (abbreviated pol I)
Ligase
Primase
• Technically, more than a dozen enzymes participate in
replication.
– Many are smaller enzymes complexed into larger ones.
• Also, the polymerases listed above were identified in
prokaryotes.
– The eukaryotic enzymes are very similar and discoveries are still
being made.
Enzymes and Energy
• Awesome. Enzymes. So?
– No, really, just because you have enzymes doesn’t
mean you can magically build a giant DNA molecule.
• And DNA is giant. Even though it’s only about two nm
wide, there is six feet of it in every cell!
– Something like DNA is highly endergonic and is quite
unlikely to happen completely spontaneously.
• So from where is our energy coming?
– Well, from where does it usually come?
ATP, GTP, CTP, and TTP
• Yep, you read that right.
• DNA polymerization uses ATP, along with its close relatives
GTP, CTP, and TTP – each corresponding to a different letter.
– In other words, the energy is packed with the raw materials.
• DNA bases arrive as nucleosides (nucleotides without the
single phosphate), and in fact have three phosphate groups
attached.
Cytidine
Triphosphate
Thymidine
Triphosphate
Guanosine
Triphosphate
Adenosine
Triphosphate
– We call them nucleoside triphosphates:
P
Formation of Phosphodiester Bonds
P
P
Deoxyribose
Thymine
 Dehydration Synthesis
P
P Deoxyribose
P
Adenine
 Dehydration Synthesis
P
Deoxyribose
Guanine
DNA Replication
• So we’ve met the “characters” (enzymes) and
the “props” (ATP, GTP, CTP, TTP).
• Now let’s watch the play…
– Heads-up: I’m going to summarize the whole
process in one big note-worthy slide first, then
we’ll look into the details of the process.
– It might be a good idea to write this with space in
between. You’ll need no more than 1/3 of a page
unless you have ginormous handwriting.
– Watch the headings to orient yourself.
DNA Replication Process
Summary Slide [enzymes underlined]
1. DNA helicase unwinds the double helix by breaking
hydrogen bonds between nitrogenous bases.
– Single-strand binding proteins prevent re-coiling.
– Topoisomerase relieves physical strain in the coiled part
of the strand.
2.
3.
4.
5.
Primase lays down an RNA nucleotide primer.
Pol III adds DNA nucleotide bases from 5’ to 3’.
Pol I replaces RNA primers with DNA nucleotides.
Ligase joins disconnected fragments of DNA.
Step 1: Unwind the DNA
• [DNA pickup line]
• For protection, the DNA
molecule is highly coiled.
– A consequence, however, is
that it also can’t be copied –
enzymes cannot access it.
• DNA helicase uncoils the
helix and creates two
replication forks (uncoiling
spots).
– Sometimes called a
“replication bubble.”
http://stream1.gifsoup.com/view3/1504259/dna-helicase-o.gif
Step 1: Unwind the DNA
• Remember that we’re
dealing with unthinking
molecules, though. How
does the DNA molecule not
just re-coil?
– Single-strand binding
proteins attach themselves
to the nucleotides to
prevent them from coming
back together.
https://dr282zn36sxxg.cloudfront.net/datastreams/fd%3A85df246e2fbc147adc76f893a37156af339c95c1a0f63df3456a110b%2BIMAGE%2BIMAGE.1 AND ALL FOLLOWING
Step 1: Unwind the DNA
• Of course, untwisting
one section of the
DNA will add strain to
the still-coiled
section.
• Topoisomerase
prevents damage to
the “upstream” part
of the strand.
Step 2: Add RNA Primers
• DNA polymerase has a major limitation:
3’
5’


– It needs a 3’ carbon to serve as a
foundation for the placement of the 5’ end.
• It’s really that little hydroxyl group that it needs
to “plug into.”
• So even though the parent DNA
molecule is ready to go, there’s no way
for pol III to start adding nucleotides.
• Luckily, there’s another enzyme out
there that can get things started:
primase.
5’
3’
Step 2: Add RNA Primers
• Primase attaches to
the DNA molecule and
adds a short stretch of
RNA to the template
parent strand (5-10
nucleotides).
• These primers provide
the 3’ carbon for pol
III.
Step 3: Pol III Polymerization
• Pol III, which is made of a bunch of subunits, starts
at the primer and adds DNA nucleotides, moving
from 5’ to 3’.
Step 3: Pol III Polymerization
• Wait…what about the 3’ to 5’ direction?
– As in, how does pol III polymerize the
complementary strand?
• Pol III does so in short stretches.
• Let’s look at this problem with a conceptual
diagram.
5’
P
Why Only 5’ to 3’?
P
P
Deoxyribose
Thymine
P
 Dehydration Synthesis
P
P Deoxyribose
P
P
5’
Deoxyribose
Adenine
Guanine
 No Dehydration Synthesis!
 Dehydration Synthesis
P
P
3’
P
Deoxyribose
Deoxyribose
Guanine
Cytosine
3’
http://a.fastcompany.net/multisite_files/fastcompany/imagecache/inline-large/inline/2013/11/3021307-inline-fb-thumbsup-printpackaging.jpg
Linking Nucleotides
• Remember this?
5’ to 3’ Confusion Point
• Look at the DNA molecule below.
• To some of you, DNA polymerase may appear to be
running backward.
– If so, it’s because you’re looking at the template strand…
– …not the daughter strand.
• Key: Always look at the daughter strand.
• Pol III reads 3’ to 5’ but
writes 5’ to 3’.
More on Replication
3’
5’
Replication
Fork
Replication
Fork
5’
3’
Note: The following slides concerning replication will feature
close-ups of different regions of the above molecule as it is
replicated, unless otherwise noted. That’s important to know.
DNA Replication: Leading Strand
For the leading strand,
primase lays down an RNA
primer (5’ to 3’).
Pol III adds DNA
nucleotides (5’ to 3’).
5’
3’
5’
3’
5’
3’
DNA Replication: Leading Strand
5’
As the replication fork
moves toward the 3’ end,
Pol III adds more
nucleotides continuously.
3’
3’
5’
5’
3’
DNA Replication: Lagging Strand
For the lagging strand,
primase lays down an RNA
primer (5’ to 3’).
Pol III adds DNA
nucleotides (5’ to 3’).
5’
3’
3’
5’
5’
3’
DNA Replication: Lagging Strand
5’
As the replication fork moves toward
the 5’ end of the daughter strand, a
new primer is needed.
We have now formed two
Okazaki fragments.
3’
5’
3’
5’
3’
DNA Replication:
Leading and Lagging Strands
Lagging
Strand
3’
5’
5’
3’
Leading
Strand
Leading
Strand
3’
5’
5’
3’
Lagging
Strand
Leading vs. Lagging Notes
Reiji Okazaki
• The leading strand is the one polymerized continuously
from 5’ to 3’ in the direction of helicase’s movement.
• The lagging strand is the one polymerized in sections.
– In the opposite direction of helicase’s movement.
– The sections are called Okazaki fragments.
• The lagging strand is still polymerized at the same
speed but takes slightly longer to finish (more on that
soon).
– It’s also still polymerized 5’ to 3’, just not continuously.
Step 4: Pol I Replaces Primers
5’
• Pol I moves in and changes all the RNA
primers to segments of DNA.
3’
5’
3’
5’
3’
Step 5: Ligase Polymerization
• Ligase joins Okazaki fragments to seal
any gaps in the DNA.
3’
5’
3’
5’
3’
Review
• Turn to a neighbor, say hello, and summarize
(without looking at your notes) the process of
replication.
– Include enzymes and directions.
Aside: Ligase Mechanism
• In case you’re wondering, ligase works by, essentially,
this mechanism:
– Ligase brings over AMP (adenosine monophosphate)
attached to an amino acid (lysine).
– AMP is attached to the 5’ phosphate of the “upstream”
nucleotide.
– The addition of AMP causes the OH- (hydroxyl) group on the
next “downstream” nucleotide to bond with a phosphate in
the upstream nucleotide.
– The adenosine molecule is then released, leaving a bond
between the 3’ OH- of one nucleotide and the phosphate of
another.
• In short, ligase “plays matchmaker” by adding an adenine nucleoside
temporarily. It then is released as the two previously separate
nucleotides decide they’re going to bond after all.
So once again, that’s…
1. DNA helicase unwinds the double helix by breaking
hydrogen bonds between nitrogenous bases.
– Single-strand binding proteins prevent re-coiling.
– Topoisomerase relieves physical strain in the coiled part
of the strand.
2.
3.
4.
5.
Primase lays down an RNA nucleotide primer.
Pol III adds DNA nucleotide bases from 5’ to 3’.
Pol I replaces RNA primers with DNA nucleotides.
Ligase joins disconnected fragments of DNA.
Or, in one image…
https://dr282zn36sxxg.cloudfront.net/datastreams/fd%3A85df246e2fbc147adc76f893a37156af339c95c1a0f63df3456a110b%2BIMAGE%2BIMAGE.1
More limitations…
• The other problem occurs at the ends of a
chromosome.
– In eukaryotes.
– Prokaryotes like bacteria have circular DNA and
thus no “ends.”
• Remember how pol III and pol I need 3’
hydroxyls off of which to build?
• Yeah…
Replication at Chromosomal Ends
The leading strand has no noticeable issue.
3’
5’
5’
3’
Primase
Pol IPol III
3’
5’
Pol I
5’
3’
End of DNA
Molecule
PolPrimase
III Pol I
The lagging strand cannot fully replicate.
At the DNA end, there is no existing 3’ -OH for pol I to use.
The Consequences
• Since pol I can’t replace the RNA primer, DNA
molecules develop staggered ends.
• This is a normal part of replication and it occurs at
the telomeres. Remember those?
– Telomeres are the “junk” end caps of a chromosome.
• They’re like the aglets (nubs) on shoelaces that prevent
unraveling.
• Interestingly, they have 100 to 1000 repeat sequences of
TTAGGG. This will come into play for telomerase soon.
3’
5’
5’
3’ Staggered End
The Consequences
• With each replication, the telomeres shorten,
which is thought to be related to aging.
– Each division eliminates between 30 and 200 base
pairs.
– The number of cell divisions that can occur before cell
cycle arrest is around 60 or so – this is known as the
Hayflick limit.
• White blood cells’ telomeres in newborns have
8000 base pairs.
• In adults, telomeres average 3000 base pairs.
• In the elderly, there are only 1500 remaining.
http://learn.genetics.utah.edu/content/chromosomes/telomeres/
Undoing Staggered Ends
• Normally, DNA with staggered ends will trigger a
checkpoint to stop mitosis.
• However, staggered ends at a telomere is normal,
so proteins at the telomeres inhibit that kind of
response.
• But what about cells like sperm and ova?
– Once it forms, it has a whole lot of divisions ahead of it
in life.
– Wouldn’t the telomeres degrade before they’re “done?”
• The answer is “no,” thanks to an enzyme called telomerase.
Telomerase
• Telomerase rebuilds telomeres to prevent
shortening.
– It extends the 3’ end so that the usual process can
extend the other side using an RNA template of
AAUCCC.
• More on the next slide.
• Are you thinking what I’m thinking?
– “If we force telomerase to become active all the
time, can we prevent aging?”
– Well…maybe.
• More in three slides.
Action of Telomerase
The telomere is shortened by a round of replication.
Telomerase rebuilds the lost nucleotides.
3’
5’
5’
3’
Telomerase
PolPrimase
III
Shortened telomere terminal
Original
Rebuilt telomere terminal
Action of Telomerase
• Telomere Replication video
Telomerase and Immortality
• Telomerase concentrations “in the real world” are highest in
cancer cells, which use the enzyme to prevent the
telomeres from shortening to the point that the cell can’t
divide anymore.
– Pancreatic, bone, prostate, bladder, lung, and kidney cancers all
feature shortened telomeres.
• In lab settings, however, telomerase has been used to keep
cells dividing continuously without causing cancer.
• Further, telomeres are not the only influence on aging, since
mice have giant telomeres compared to much longer-lived
humans.
– Let’s hear more from the voice of Richard Cawthon at the
University of Utah.
http://learn.genetics.utah.edu/content/chromosomes/telomeres/
Aside: Dyskeratosis
• Dyskeratosis is a disease that causes premature
telomere shortening.
– The result? People with the disease age and die much
more quickly than those without the disease.
– Other symptoms:
•
•
•
•
•
Early hair graying and balding.
High leukemia risk.
High cirrhosis risk.
Learning disabilities.
Bone softening.
http://learn.genetics.utah.edu/content/chromosomes/telomeres/
DNA Can’t Spell
• Okay, that’s a little harsh. DNA actually spells
pretty well most of the time, but every once in
a while it puts a letter or two in that shouldn’t
be there.
• As you might guess, evolution also brings us a
fair degree of typo recognition and error
correction.
– Naturally, some slip through even these layers, but
at least it’s something.
• As you also might guess, it’s enzyme-driven.
Polymerization Speed
• Pol III runs at about 1000 bases per second as the
main DNA builder.
• Pol I runs at about 20 bases per second.
• Why the speed difference?
– Pol I is actually a proofreader, too!
• We know Pol I removes primers, but it also does
some error correction (known as mismatch repair).
– The error rate without pol I is 1 in every 10,000 bases.
– The error rate with pol I is 1 in every 100,000,000 bases.
How it works…
• Another enzyme, called a
nuclease, literally cuts the
erroneous nucleotides
out.
• Pol I then replaces the
DNA with appropriate
nucleotides.
• Ligase, as usual, steps in to
seal up the strand.
• Fun fact: Pol II appears to be
involved in error checking in
prokaryotes.
Mismatch Repair/Action of Nuclease
Damaged DNA is excised by nuclease.
Pol I and ligase proceed as normal.
Nuclease
3’
5’
5’
3’
NucleasePol
I
Ligase
Remember this?
http://skreened.com/scienceforscientists/dna-checks-itself-before-it-wrecks-itself
Closure
•
•
•
•
•
So…DNA.
It’s the stuff of life, right?
Worth another 100+ slides, right?
Why?
What does DNA do for you?
Closure
• DNA can also be used in an “outside the body
sense.”
– I mean like in a “forensics” sort of way.
• Especially since a lot of DNA in forensics is outside the
body.
• Still other uses come from forensics of other
creatures.
• No, not “it was the giraffe in the study with the lead pipe.”
– I mean like in a “clues in DNA that tell us about the
evolutionary relationships between organisms.”
Closure Part Deux [Videos]
•
•
•
•
•
DNA Replication Fork 1 animation
DNA Replication Fork 2 animation
Honors DNA Replication 1
Honors DNA Replication 2
DNA Replication 2 – Molecular – Advanced
Closure Part Three
• Turn to your neighbor, say hello, and take turns
narrating the steps of DNA replication.
– Include error-checking this time.
• For example, suppose you and your partner are
talking:
– James Westfall: “Hello.”
– Dr. Kenneth Noisewater: “Hey there.”
– James Westfall: “So, DNA replication starts with
helicase.”
– Dr. Kenneth Noisewater: “Helicase works by…”