[ Silence ]
>> My name is Javert.
Was that at least as
good as Russell Crowe?
OK, so we're ready
to get started.
First a quick quote,
"Chemistry is to biology,
what notation is to music."
To me this really grabs at the
essence of chemical biology
in the sense that the notations
on a musical scale
allow creativity.
They allow other reformers to
interpret the works in new ways
and give the work context.
Chemistry does that in biology.
Chemistry gives us
an opportunity for us
to be creative about
biology and invent new ways
of thinking about biology.
It's sort of the underlying
basis at the level of atoms
and bonds as I keep
saying, for biology,
and to me in some way this
really captures what we're
trying to do in this class.
OK. So this week, where
it's already week 3
which is amazing-- Oh, hang on.
OK, so it's week 3 so we're up
to chapter 3 and we're going
to be talking about DNA.
Our knowledge of DNA
was really set in place
by the people in front of you.
These are the giants really in
the field of structural biology
who determined structures
of DNA in the 1950s.
This includes the great Rosalind
Franklin whose very accurate
x-ray diffraction
structures and her pictures
of the x-rays diffracting
off the fibers of DNA set
in motion the determination
of the structure.
She was working with Maurice
Wilkins and two physicists,
Francis Crick and
Jim Watson went
on to solve the structure
of DNA.
And as we'll see in a moment,
really one of their key insights
was at the level of atoms
and bonds in the sense that
they discovered interesting
tautomerization of the DNA
bases that made it possible
to have what we now call
Watson-Crick base pairing
between the strands of DNA.
Getting a little ahead of myself
but that's where we're going
in the next week or so.
So we're going to be finishing
up non-covalent interactions
then talking
about DNA structure,
DNA property
and finally DNA reactivity
of small molecules.
This is a large chapter.
We have a lot to talk
about, so bear with me.
Things are going to go
not faster it's going
to be the same speed,
but we're going to gloss
through a few topics that are
less important and when we do,
this means then that you
can focus your reading
and your study just
on the level of detail
that we're covering
in the class.
OK. Some announcements, in
the textbook, read chapter 3.
Again, skim concepts not
presented in lecture,
don't get too worked
up about them.
Chapter 3 problems, do all
of the odd numbered and all
of the asterisked problems.
In addition, I want
to encourage you
to get involved here
at UC Irvine.
This is super important.
Many of you I know aspire to
become physicians or scientists
or pharmacists or whatever
it is that you aspire to do.
All those big plans
require preparation,
they require some evidence that
you've gone beyond the ordinary
and I want to encourage
you to do this.
OK. One way to get
involved is to look
around for opportunities
to volunteer.
This is one that's run
by my friend who is one
of the founders, it's called
the Social Assistance Program
for Vietnam.
If you go to this website, there
are opportunities to volunteer
to spend two weeks in
Vietnam, in a rural part
of Vietnam administering
medicine, you know.
You'll probably not be
of course, you know,
drilling people's teeth and, you
know, doing open heart surgery
but you will get a
unique opportunity
to actually see those
types of things happening
and that's really
important if you aspire
to that kind of career.
It provides evidence that you're
qualified, that you're committed
and that you're someone
who is altruistic.
All of those things professional
and graduate schools look
for in your application.
You need to be doing
those things now.
OK. And I'm on your
side on this.
OK. I will help you get--
find those opportunities.
I'll bring them to your
attention like this one.
And if there's something
in particular that I can do
to connect you with, let me know
and I'll do my very
best on your behalf.
OK. Along those lines, our
laboratory always has openings
for talented undergraduates.
It's competitive but you
have a chance to participate
at the full level of
a graduate student.
Undergraduates in our laboratory
are doing actual science.
They're publishing
papers with us.
They're making discoveries
and they're participating
as full members of the team.
OK. Here's how you apply,
send me a paragraph
describing your career goals
and how research
in our laboratory would
advance those career
and educational goals.
In addition, send me a copy of
your college level transcripts.
This includes any transcripts
at community colleges
if you're transferring.
Many of my best students
are transfer students.
Send me those transcripts
as well
and also send me three names
and email addresses of TAs
who know you well
in lab sections.
OK? And I'm going to
email them and I'm going
to ask them what was this
person like in the laboratory?
Were they, you know, the
first one out of the room?
Were they last one
out of the room?
Did they, you know, follow you
around the laboratory
asking you,
"Does this look pink,
does this look pink?"
or were they pretty
independent, OK?
So I'll find out about
that sort of thing
and then that's how I
make a decision on who
to accept into the laboratory.
OK. And then of course
the resume.
This is pretty standard,
if you're interested
in doing research here at
UC Irvine which I highly,
highly encourage you to do,
this is a pretty good
way to go about it.
OK. This is an effective way
to get noticed and to get
that job that you need.
OK. Any questions about
these opportunities?
Why I think they're important
and things like that?
OK. See me in office hours if
there's something in particular
that you want from me and
I'll try to hook you up.
OK, office hours this
week, speaking of which.
Tomorrow I'll have
my usual office hour,
2.45 to 3.45, the
usual location.
Thursday I'll have my office
hour 11 to 1, usual location.
In addition, Mariam will have
her office hour Fridays 1:30
to 2.30.
And, Kritika, could
you raise your hand?
So, Kritika is our new TA.
She'll be joining in the team.
And Kritika, does this time work
for you, Tuesdays 2:30 to 3:30?
Good. OK. And she'll be having
her office hours Tuesday.
So notice that we spread
out our office hours
so that there's one everyday
of the week except Monday
because I know you're very busy
on Monday doing all
kinds of things.
I hope you're having
fun yesterday.
But yeah, so everyday of the
week there's an office hour,
they're staggered so
they're at different times,
so you can have your
questions answered.
And again, Kritika is a graduate
student in my laboratory.
She knows this material
as well as I do.
She's really smart.
You can go to her office
hour and get an answer.
That's as good as an
answer that I will give you.
OK? And for that matter
you can also email the TAs
with your abundant questions.
OK. I'm looking at you where
I can find that person, OK,
there's like one
person in the class,
he send me 10 emails a day but,
you know, I will do my best
but you can also
email the TAs as well.
OK. Oh, along those lines,
I sent you an email
saying don't send me book
or potential journal articles.
And the reason is I must do-- I
open my inbox and I had like 15
of those and I got to the point
where I was bouncing messages
because the inbox was so full.
So, if you send me those, I
can't do very much with them,
OK, it might clog my box.
So what I propose we do is
instead of you emailing me them,
instead bring them
to my office hours,
bring them to Kritika's office
hours or Mariam's office hours
and ask your questions then.
OK. Now the standard
question I get asked is,
is this article appropriate?
And my answer to that is if
you follow the guidelines,
it will be appropriate.
Now in addition, when
you're writing your summary,
your report on the journal
article, focus on the aspects
of the article that fit the
definition of chemical biology.
OK. So a paper in cell
for example is going
to be a very meaty paper, it's
going to cover about 10 pages.
It's going to have, you know,
eight or nine figures and some
of those figures, some of those
experiments aren't exactly what
we will call chemical biology.
Don't focus on those.
Focus on the ones that are
chemical biology related.
OK. Otherwise I don't know
that you know the definition
of chemical biology.
OK. Any questions?
All right, guess what, we're
heading into midterm season.
This is week 3, so
week 4 is next week.
We will have a midterm
next Thursday,
a week from this Thursday.
There will be a review
session in advance
by the TAs, time
to be announced.
Kritika will arrange for this.
The seating for the
midterm will be assigned.
Mariam, you'll do
the assignment.
It's really essential that
you bring your UCI student ID.
We will check the IDs to
make sure you're seated
in the right seat.
If you're not seated in the
right seat, it will be treated
as an academic honesty
infraction.
No notes, no calculators,
no electronic devices,
you don't need them.
You're smart.
OK? Any questions about
the upcoming midterm?
OK, now I know you want to know
what will be on the midterm.
OK, so let me tell
you, it will cover
through Tuesday's lecture
one week from today
and so we will be about halfway
through chapter 4
on Tuesday, OK?
So plan to read through about
halfway through chapter 4,
that's the chapter on RNA and
that's where I expect to be
for Tuesday's lecture.
It's possible I might
get behind but I'm going
to really try hard
not to do that.
OK. All right, I will also
post a practice midterm
to the website and
you can use that along
with the discussion worksheets,
the assigned problems as a guide
for what will be on the midterm.
OK? So the midterm will look
very much like a compilation
of discussion worksheets,
of assigned problems
and the practice midterm.
OK. And it will be about as long
as the practice midterm as well.
So when the practice
midterm comes out,
I'll post two versions.
One version will be blank,
one version will be the key.
The blank version
you should print out
and then give yourself an hour
and 20 minutes and make sure
that you can handle it.
OK. And at the end of that,
then check your answers
against the key.
But give yourself
a real practice.
OK. That's pretty
important, I think.
OK, so anyway, that's
the plan, any questions
about the midterm coming up?
I know you will have
lots of questions.
I look forward to hearing
about them in my office hours.
All right, I want to go back
to finish up our discussion
of non-covalent interactions and
where we left off last time was
with charge-charge interactions.
I'm not ready to talk to
you about interactions
between atoms that
are uncharged.
OK. Neutral atoms that are
interacting with each other.
These are described by a
Lennard-Jones Potential
which is an equation
that describes how these
neutral atoms would interact
with each other.
Another way of describing
these neutral atoms,
another term that's
used and probably one
that you encountered is
London dispersion force.
OK. So when you have two say
neon atoms that cozy up next
to each other, then
they will interact
through a London
dispersion potential or force
and that's what I'm
describing here.
OK. So, it's just a
couple of different ways
of saying the same thing.
This happens a lot in
biology, not necessarily
between neon atoms but certainly
between aliphatic side chains,
hydrophobic side chains in
proteins, in interactions
with each other, interactions
with lipids, at the--
a plasma membrane of the
cell, and a whole host
of other non-covalent
hydrophobic-hydrophobic
type interactions.
This turns out to
be a very potent
and very strong force
in biology.
OK. So we need to
understand it better.
So the energy-- the
potential energy
of a van der Waals interaction,
yet another word to describe it,
is equal to-- is
proportional to 1 over r
to the 12th minus
1 over r to the 6.
These terms, the sigma term
deals with the diameter
in this epsilon ij, not so
important, so let's ignore that.
Let's focus in on the 1 over
r to the 12th term and 1
over r to the 6th term.
First, notice that it's minus
1 over r to the 6 and minus
in potential terms means
more stable in energy,
lower on this Y-axis of
potential energy over here.
OK. So that's going to
be our attractive term.
Hydrophobic, if things attract
each other, OK, not just due
to repulsion from water.
We'll talk about that next.
But hydrophobic things
want to stick to each other
and they're going to do this
with an attraction
that's proportional
to 1 over r to the 6.
The fact that it's 1 over r to
the 6 as opposed to r to the 2nd
in the charged-charged
interactions means
that this is a much
shorter range attraction.
This attraction takes place
on a very tiny distance scale.
OK. Now eventually the two atoms
in this case as described here,
or two molecules-- or two
molecules bang into each other
and go past the point where
they're attracted to each other.
OK? And at that point,
their electrons are trying
to overlap with each other.
That's really bad news, right?
We know by the Pauli
exclusion principle
that that's not allowed
and so in the same way
that my fingers are never
going to fuse with each other,
just going to bang off of each
other, the atoms push away
from each other and they
push away from each other
with the repulsion force
or repulsion potential
that's proportional
to 1 over r to the 12th.
OK? And so this means that
this is extremely short ranged
and extremely sharp, right?
To the 12th power
is a large number.
So this means that this really
dramatically pushes apart the
atoms if they happen to get
too close to each other.
It turns out that there
are a whole series
of other non-covalent
interactions that we find
in biology that actually
contribute quite a bit
of non-covalent binding energy.
Here for example are the
dispersion interactions
that we have discussed
before on the previous slide
and so these include things
like aliphatic-aliphatic
interactions,
but also aliphatic interacting
with hydrophilic molecules.
So here is water
interacting with methane.
They're going to
interact with each other
and have some attraction.
These number here, minus 0.5
to minus 0.7 kcals per
mole is pretty low.
OK. This is not a
tremendously strong interaction.
Where it gets strong is
when you have a molecule
that has a large number
of functional groups.
Each one with 0.5 kcals per
mole here, 0.5 here, 0.5 here,
and when you sum up across
all of these, you're starting
to talk about big energy.
OK. Now just to give you an
idea, you need to know one fact
that I think is really
important.
And the fact is important
enough that I'm going to try
to write it on the board
over here in the corner.
The fact is that, a factor
of 1.4 kcals per mole will
be a factor of 10 in--
at equilibrium constants.
OK. So, 1.4 kcals per
mole is a magic number
in chemical biology.
OK. So look for 1.4
kcals per mole
because that tells you
then that's favored tenfold
over nonbinding.
In other words, the
interaction is going
to be 10 times more likely
to form than not form.
OK? It's a factor of 10 in
terms of equilibrium constants.
OK. So, if we're talking
about something over here,
that's only 0.5, 0.7 kcals per
mole, you have to start summing
up a whole bunch of these
to get anywhere in terms
of enforcing the interaction.
On the other hand, some of these
other interactions can be quite
strong and let's take a
closer look at those next.
OK. So, for example,
we've talked a little bit
about hydrogen bonding.
Hydrogen bonding of course
has a donor and acceptor
and here's a range of strengths.
Hydrogen bonds vary
enormously strength
from about 1 kcal per mole all
the way to 7 kcals per mole.
The strength of the
interaction depends enormously
on the identity of the
donor and acceptor.
When the donor or-- and/or
acceptors are charged,
if either one is a charged
functionality, the strength
of this hydrogen bond
goes up enormously.
And this kind of
makes sense, right,
because remember earlier I
described a hydrogen bond
as a kind of a special case of
a charged-charged interaction
in which a hydrogen is being
shared between two atoms.
OK? So, if one of these happens
to be charged, that's going
to be a much stronger
charged-charged interaction.
Speaking of charged-charged
interaction,
salt bridges are the
coulombic potential
that we saw on Thursday.
These are the charged-charged
interactions.
These vary also enormously
depending upon the environment
that the salt bridge happens
to find itself in, where a--
where water can shield
this charge.
Water or counter ions
can shield this charge,
decreasing it considerably
and making the interaction
much, much weaker.
So, a salt bridging interaction,
which is another way
of saying charged-charged
interaction found
in a hydrophobic
environment, say the interior
of a plasma membrane, is going
to be a much stronger
interaction
than one that's found out in
water where there's plenty
of water and counter ions
to shield the charge.
OK, where those provide a
counter against the charge.
Recall that those environmental
terms are embodied by the 1
over 4pi epsilon term in
the coulombic potential
that I showed you on Thursday.
OK. In addition, there's also
dipole-dipole interactions
which are alignments
of densities of charge
where we have a little
bit more negative charge
on the oxygen over here.
The dipole is pointing
in this way on the--
to the right on the
upper acetone
and to the left on
the lower acetone.
The two of these dipoles want
to cancel each other out.
By cancelling each other out,
that will give you a more
optimal interaction and that's
where some potential energy.
Finally, there's also a
whole series of aromatic
or arene interactions.
And in general, this includes
both face-to-face interactions
where you have two
faces of a benzene ring
that are interacting
with each other.
Notice in this picture over here
that the top benzene
ring is offset
from the bottom one
and this makes sense.
We're going to be
looking at regions
of electron density interacting
with regions of electron
poverty.
OK, that that's actually the
basis for the interaction.
And so for that reason, we
also see very commonly edge
to face interactions.
OK, so this is the one
that we'll see in a moment
when we start looking
at pi stacking in DNA.
But in addition,
you can have an edge
of an aromatic system
interacting with the face
of another aromatic system down
here, and that's as strong,
right, it has the
equivalent strength.
Even though you expect, you
know, face-face to be ideal,
that's actually not what we
see when we start looking
at large numbers of
aromatic interactions.
We see this H-- edge to face
interactions all of the time.
OK. And then finally
there's some other ones
that are really bizarre and
they include charged interacting
with the electron
rich aromatic rings.
And this kind of
makes sense, right?
You have something
that's positively charged,
you have something
that's very electron rich
in terms of the ring system.
So these cation-pi interactions
which is what this one is called
are found pretty ubiquitously
in biology, oftentimes
playing a commanding role,
playing a really key
role in chemical biology.
OK. So, these are ones that
I'd like you to memorize.
I'd like you to know something
about their strengths,
which one is strong,
which one is weak.
I don't want you to memorize the
numbers per se but I want you
to know something
and be conversant
on relative strengths.
OK, relative strengths matters.
OK. And one thing-- one
last thing to keep in mind
if we're going for this
1.4 kcal per mole, again,
you can have a summation of a
large number of interactions
to achieve that 1.4 kcals
per mole or even more.
And I'll show you an
example of that very shortly.
Now it turns out that it's
actually a little bit tricky
to start comparing
energetics when you design
in say the perfect
cation-pi interaction.
What ends up happening is
that you get a complication
due to water, OK?
So let's imagine
that you had designed
in the perfect cation-pi
interaction and in doing
so you put this positively
charged thing that forces all
of the water around it
to rearrange itself
or reorient itself.
It turns out that's
actually a complicated thing
of the orient-- reorientation
of water
but it cannot be neglected.
OK. So what we do is we make
a very important simplifying
assumption, and I'll talk more
about water on the next slide.
But before I do, water, since
we just have to acknowledge
in advance, water can
complicate everything, right?
It's present at 55 molar
concentration in your cells
and we can't neglect it, OK.
It has its own energetics.
It's-- as I showed on
this slide over here,
for example it's interacting
with hydrophobic things.
So its own energetics are
really complicated, OK,
and actually very hard for us
to understand and pin down.
And so it's really difficult
to estimate the entropy lost
or gained in an interaction due
to that rearrangement of water
when you start making changes.
So what we'd like to do
is compare things that are
as similar to each
other as possible.
OK. This is the simplifying
assumption
that I alluded to earlier.
Here for example is an
example of that, OK?
So, here's two possible
transitions states,
and transitions are two
possible mechanisms.
Mechanism number one
involves an SN2 reaction.
Mechanism number two involves
the same molecule undergoing an
E2 elimination reaction.
And the key here is that
the molecules are identical.
OK? That extreme similarity
makes the comparison
between these two
much easier to make.
OK. And so, for example if
we're looking at two proteins,
we can look at empty protein
versus ligand-bound protein.
But on the other hand, we're
not trying to make all kinds
of changes to the protein
structure over here.
Problem is proteins are rarely,
you know, like looking like this
when the ligand is unbound.
So, these simplifying
assumptions will start
to cause all kinds of problems.
Here's one though that works.
You can make a single change
to the surface of a protein
and then compare
the altered protein,
compare its interaction
with a ligand.
So for example, we could
change this isopropyl group
to a methyl group and then
compare what's happened,
what's different in that
receptor ligand interaction, OK?
So all you've done is to
remove two methyl groups.
That's about as simple
as it gets, right?
So that type of experiment is an
easy one to make comparisons to.
OK. And again, by doing
that, we're trying
to minimize how much
the water has
to rearrange itself
at that interface.
OK. It turns out actually
this assumption works most
of the time.
So in short, being good
scientist, not changing lots
of variables at the same
time pays off in biology
because underlying everything we
do is this complicated solvent
that we operate in called water.
Let's take a closer look
at the structure of water.
OK, so here is water in ice and
notice how neatly regular it is
and how nicely ordered it is.
And then here's water in a
solution as liquid water.
And it's just crazy complicated.
First, notice that
there, all these dots--
dotted lines are
the hydrogen bonds.
These hydrogen bonds are
pretty much maximized.
Water is not passing
up any opportunities
to hydrogen and bond to itself.
OK, but the hydrogen bonds
in the liquid solution
are nonoptimal.
OK, water in solution, each
water molecule is jam-packed
with other water molecules
and oftentimes the hydrogen
bonds are slightly distorted
or they don't have
the right distances.
Those little distortions
and that lack
of perfect distances
makes the hydrogen bonds
in liquid water weaker than
they are in solid water.
Furthermore, a molecule of
water in its own, you know,
with a lot of other
molecules of water is--
and behaving kind of like it's
on a crowded dance floor, OK?
So, it's bouncing around wildly
against these other, you know,
molecules that are nearby and
interacting with lots and lots
of different molecules nearby,
constantly breaking interactions
and forming new ones.
OK. So, water is
actually very complex.
Weak and distorted
hydrogen bonds, OK.
In addition, when water cozies
up to hydrophobic surfaces,
it tends to form a very
ordered structure that starts
to look a lot like the
structure found in ice.
And this works by water form--
satisfies its propensity
formed hydrogen bonds
by forming a clathrate-like
structure.
So for example, here is a
molecule of methane encapsulated
in one of these clathrates
of water,
where clathrate is just
simply a structure of water
that satisfies its desire to
form hydrogen bonds with itself.
OK, or with other
molecules of like kind, OK.
This really dramatically
changes the strengths
of nearby non-covalent
interactions, OK.
This does things to strengthen
those non-covalent interactions
because every time one of those,
let's just say
hydrophobic-hydrophobic
interactions breaks,
then water has to slide
in between the now
broken interaction
and form one of these
clathrates.
OK, that formation of the
clathrate, the formation
of an ordered structure
cost energy.
It's a loss of entropy.
This is a more ordered
structure than the structure
of disorganized water
that I showed you
earlier in solution, OK?
So for this reason,
hydrophobic--
hydrophobic molecules are
driven against each other.
They want to find
each other in water.
And this is sometimes
referred to as a-- this is--
this is actually a
water driven effect.
Forgetting the technical
word for this.
OK, anyway, so--
oh sorry, it's--
it's sometimes referred to
as a hydrophobic effect,
OK, in water.
OK, now let's take a closer look
at a receptor-ligand
interaction,
now zooming in at the
level of atoms and bonds.
This is a molecule called
human growth hormone and, yes,
Lance Armstrong admitted
to Oprah
that he took human
growth hormone to win--
to help him recover basically
from different stages
of the Tour De France during
all seven of his victories,
and it really annoys
me actually.
I could say a lot
more about that
but I'm going to
hold myself back.
OK, now when human growth
hormone binds to its receptor
on the surface of cells, it's
stimulating growth and recovery
of those cells, it's stimulating
protein production, et cetera.
And when it binds to
the surface of the cell
to the binding partner on
the surface of the cell,
its receptor, then all of
the region that's colored
in on this surface
is buried, OK?
So in other words, human growth
hormone binding protein binds
over here and then
makes contact with each
of these colored atoms.
OK, everything that's in white
here is still out in the water,
out in the solvent,
it's not interacting
with the receptor at all.
Now, when I was at postdoc, I
repeated a classic experiment
that was done by Jim Wells.
And Jim Wells and
his co-workers found
that even though there are
19 residues that are buried
on the surface, there are 19
amino acids that are buried,
only the ones in red are
actually contributing binding
energy, OK?
So, notice that all of
these other stuff is in--
that is in blue that
is buried is not
at all contributing any
binding interaction.
So although there's interactions
between these side chains
of these two proteins, there is
no binding energy that's being
exchanged or gained
by that interaction.
OK? So, just because two
molecules find each other,
two functional groups find each
other in space, does not ensure
that there's actually going to
be a net gain in binding energy.
Because again, that net gain
in binding energy includes both
the strength of interaction
but must also include the water
ordering and disordering term
which we've been calling
entropy earlier, OK?
So, in order for this
interaction to take place,
you're going to be
pushing out ordered water
and gaining some
entropy in some places
and in other places
losing some entropy.
OK, now when we look even
more closely, let's just zoom
in on this red patch over here.
This red patch has been termed
the hot spot of binding energy.
That's where the binding energy
allowing these two molecules
to interact with
each other is found.
OK, this is the essence of
the non-covalent interaction
between human growth hormone
and its binding partner,
human growth hormone
binding protein.
And in green, these are
the functional groups
that are found in
this red patch.
OK? So the red patch is over
here and now I'm showing you
that the functional
groups were in green.
These are carbon atoms.
In blue, that's a nitrogen.
And in red, that's an oxygen.
OK, notice that the hydrophilic
functionalities, the guanine--
of an arginine over here,
a bunch of nitrogens,
another nitrogen over here, an
oxygen, an oxygen over here.
Notice that those are around the
periphery of this red region.
They're around the outside of
this hotspot of binding energy.
The center of the hot spot
is largely hydrophobic, OK?
Notice that it has lots
and lots of carbons.
There's a benzene ring,
it is smack in the center.
There's this aliphatic
chain that's capped
by an amine functionality,
but nevertheless this
is an aliphatic chain.
There's aliphatic
functionalities over here
and over here and
over here, et cetera.
OK, so in other words,
the outside hydrophilic,
the inside hydrophobic.
And so, when molecules,
functional molecules find each
other, this is a very common way
for them to interact with
each other through a small set
of residues that form this
hot spot of binding energy
which again kind of looks like
a core sample through a protein.
Outside is hydrophilic,
inside hydrophobic, OK?
Any questions so far?
OK, let's talk one last--
about one last section
of chapter 2 before we
move on to chapter 3.
There's this concept
that the biooligomers
on earth are highly modular.
We've discussed this before.
This also extends to the
polyketides and the terpenes
which are composed of
isoprenes and the polyketides
which are caused-- which are
composed of either malonyl
or acetyl subunits that
are strung together,
where the red bonds indicate
that where the connection
between these modules
such as the amino acids
as individual modules
in a protein.
OK? And furthermore, this is
also found in oligosaccharides
where you have this
glycosidic bond
that connects the glycan
fragments together.
There's also a numerical
amplification and biosynthesis.
So, if there's only
one or two copies
of DNA per cell depending upon
whether it's a prokaryotic cell
or eukaryotic cell,
some prokaryotic cells are
[inaudible] more than one
but let's just simplify it.
Then to RNA, each DNA is
transcribed 10 to 50 times
and then each RNA is
translated say 10 to 20 times.
So in the end you end up with
this massive amplification
signal going through the
cell, where with one copy
of DNA you can end up
with millions of products
from some enzyme
reaction down here.
Last thoughts, form
follows function in biology.
These-- the bonds
that join together,
the oligomeric subunits are--
have a strength that
follows their function,
their functional
requirements, OK.
And so, for example, when we
look at the half-life of lipids,
we find that actually the ester
bonds in a lipid have a halfway
on the order of a year or so.
OK, so esters, not so stable.
Compare that against DNA down
here which has a half life
on the order of 220
million years.
OK, that's his half
life for DNA.
And in retrospect, this
kind of makes sense, right,
because DNA has to be a-- you
know, has to be a biooligomer
for the life of the
organism, OK?
And so, we're now at the point
where we're routinely
taking advantage
of this tremendous stability
of DNA to amplify DNA
from even extinct organisms like
wooly mammoths, like species
of prototypical humans that
haven't lived on the planet
for tens of thousands of years.
That sort of thing
is going on right now
in laboratories talking
advantage
of the tremendous
stability of DNA.
Now your hair, which is
a protein, has a lifetime
on the order of, you
know, 300 years or so.
And you can see that, right?
We can find, you know--
we give-- well, anyway.
So, I guess it depends on the
human that we're talking about.
My hair obviously
doesn't exist that long.
But, you know, so certainly the
lifetimes here are following
their function, right?
Proteins don't have
to last this long.
Question? How does one get a PhD
that's going to take you five
or six years studying and trying
to measure these half
lives of 220 million years?
Anyone have any ideas how
to do that experiment?
I can guarantee it to you,
it's not like, you know,
you set up this test tube
and then you check it
every 20 years, OK,
to see how much gets cleaved.
How would you do this?
Yeah, how would you do it?
[ Inaudible Response ]
OK, a small amount of RNA.
And I would use a large amount
because very little is
going to get degraded.
How would you do this though?
Yeah?
>> Maybe we can put it in a
very decomposing environment?
>> OK. But then you
wouldn't know
if the decomposing
environment is different
than in the cell, right?
We want to know about the
half lives in the cell, right?
Yeah?
>> Use a model organism.
>> Model organism.
No, I want to know
what it's going to be--
what's going to last in,
you know, in this cell
or this other one over here.
Question over here?
[ Inaudible Question ]
OK, you're definitely
going to use radioactivity
because you need something
that's super sensitive.
How would you do this?
[ Inaudible Response ]
OK, you're getting close.
What is your name?
>> Bryan.
>> Bryan? OK, Bryan
is getting close.
So the suggestion
was radioactivity.
Bryan's suggestion is you look
for a tiny little quantity
and radioactivity gives
you that sensitivity.
But are you going to do this
for 220,000 years or
220 million years?
>> No.
>> OK, so how are we going
to do this experiment?
We have the sensitivity,
we're going to look
for tiny little quantities
and extrapolate back.
How are you going to
model 220,000 years?
Yeah?
[ Inaudible Response ]
Carl, OK, look at fossils.
Yeah, and we do that.
Yeah?
[ Inaudible Response ]
OK C14.
[ Inaudible Response ]
OK.
Yes.
>> You would compare it to that.
You note the reactivity of
one and then you compare it
to the one that you know.
>> Ahuh.
>> And it is-- and
extrapolate that half life,
half life, half, half, half.
>> OK, so-- but the problem
is you wouldn't know all the
conditions that's
experienced over, you know,
say 100,000 years
or something, right?
So, I mean, how do you--
you want to do this in a
controlled circumstance.
You want to have everything
just in a little test tube
where you know exactly
what's been added
to the test tube, right?
But you don't want to wait
around for 220,000 years
or 20 million years,
what are you going to do?
OK, I'd like you to
look this one up.
This is one that you
should be able to design.
Look it up.
And then when we come back on
Thursday we'll talk about this.
But I'd like everyone to
have to look this up, OK?
This is important.
OK, let's talk--
let's summarized what we've
been talking about in terms
of non-covalent interactions.
These are completely
ubiquitous in biology.
Good news, we only have
to learn two equations
which govern all interactions
in chemical biology.
Those were the Coulomb's law for
the charged-charged interactions
and the Lennard-Jones potential
for the uncharged
interactions, OK?
And so if we know those
two equations, we're set.
What's really-- and what's
important to us is not
that we're going to be plugging
in, you know, charge of this
and then, you know,
radius of this.
What's important to us
are the relationships,
the distance dependence,
the 1 over r squared
versus 1 over r 6.
That type of distance dependence
makes a big difference.
And knowing that sort
of thing and having sort
of an intuitive grasp of that
is going to be very important.
So-- and I'll just give
you a quick example.
For example, we now know if
DNA is negatively charged,
it's going to attract
other charged ions to it
from great distances, right?
Because it's distance
dependent, it's only 1 over r
to the 2nd power
versus1 over R to the 6.
In addition, we've learned
that these non-covalent
interactions are very sensitive
to the environment, the
distance and the geometry.
Water is a really
slippery molecule
to understand, to say the least.
Has a malleable structure
and it can dramatically
alter the strength
of non-covalent interactions.
This makes it really tough for
us to draw any generalities
because water is an intermediate
lubricant between all
of these interactions and
it plays a complicated
and sometimes hard to us-- it's
hard for us to define role.
And there's still big
arguments that are going
on in water chemistry
to this day.
For example, there's
an argument going
on about how many ions are
found on the surface of water
or what's the pH at
the surface of water.
And there's been a
set of dueling papers
that have appeared that
contradict each other.
The first paper had a title
like the pH of the surface
of water is more acidic.
The next article--
the next article
by the competitor
said the pH of water
at the surface is more basic.
And the two-- and these groups
have been arguing backward
and forth and both making very
reasonable arguments for years.
OK? The truth is what we found
is actually it's somewhere
in between these two and you
can actually see evidence
for either one and it turns
out to be a very minor
effect that's not
so important in biology.
But the point is is
that water itself is
such a complicated fluid
that we're still using the
latest techniques to try
to understand it better.
It's not fully understood.
Hydrogen bonds have
donors and acceptors
and they are also very
susceptible to a competition
with water for those
hydrogen bonds.
I would like you to know
the approximate strengths,
the relative strengths,
not the approximate
but the relative strengths,
and distance dependence
of non-covalent interactions.
That's important.
OK, so that's a summary
of chapter 2.
Any questions about chapter 2?
Yes, Chelsea.
[ Inaudible Question ]
Yeah, I really want
you to know that.
OK, that's super important.
That's that
Henderson-Hasselbalch equation.
That hopefully you
learned in Chem 1,
you definitely need
to know that.
Other questions?
OK, let's move on.
I want to talk to you
about the structure of DNA.
This is the classic structure
of DNA first proposed by Watson
and Crick in I believe 1952 or--
yeah, 1952, somewhere in there.
The structure of DNA
has two strands running
in opposite directions
to each other.
So they're anti-parallel
to each other.
The strands are held together
by phosphodiester bonds
which we'll look
at more closely.
So, here's a schematic
diagram of what the structure
of DNA looks like and
here's a space filling view
where each one of these spheres
is a van der Waals' sphere
to approximate where
the atoms are,
where the outermost
electrons of the atoms are.
One thing to notice is that
DNA has two grooves, OK?
It has, yeah, the distance here
between these two
strands is very close
versus the distance here between
the two strands being much
further away.
These are going to
be called the minor
and major grooves respectively.
And this is the origin of the
fact that DNA is a double helix.
I think it's commonly thought
that DNA is a double helix
because it's two relatively
rod-shaped molecules
that are twisted
with each other.
But that's actually
not the case.
It's a double helix because
it has a minor groove
and a major groove.
And I believe the next slide
will show us that more closely.
OK. So, in blue, this is
the major groove of DNA,
and in green, this
is the minor groove.
In red, this is the
phosphodiester backbone of DNA
that we've seen before, OK?
So again, notice that
there are two helices
that are running parallel to
each other, a major groove
and a minor groove, OK?
The structure of the bases
is going to set up this major
and minor groove relationship.
As we will see shortly, DNA
bases, base pairs form a U shape
and that U shape ensures that
you're going to get a major
and a minor groove, where
the inside of the U is going
to be this minor groove
and the outside will be the
major groove.
But I'm getting a little
bit ahead of myself.
The reason why this is important
is as we'll see in a moment,
proteins like to interact
with the major groove of DNA,
whereas they can't fit in to
the much closer interstices
of the minor groove of DNA.
Rather small molecules will fit
into this minor groove and try
to largely avoid the less
cozy major groove of DNA, OK.
So, almost immediately we can
start to make some predictions
about where stuff
binds just knowing
that DNA is a double helix,
double by virtue of the fact
that it has two parallel
helices,
minor and a major groove.
So, this DNA structure
immediately sets up replication.
This is the original 1953
paper by Watson and Crick,
and this is the very last
sentence of the paper
in which they had this
incandescent understatement.
It has not escaped
our attention,
it has not escaped our notice
that the specific pairing we
postulated immediately suggest a
possible copying mechanism
for the genetic material, OK?
So, if you have two strands
of DNA running anti-parallel
to each other, you can simply
separate out the two strands
and then get a perfect copy
of one strand over here
and a perfect copy of the
second strand over here, OK?
So, here's the parent
strand of DNA and again,
here are the two new
strands in orange and blue.
Note too that DNA forms
a right-handed helix.
OK, does everyone see that you
can trace out along the right--
with your right hand over
here the structure of DNA?
I think it's worth trying that.
Whereas your left hand
kind of slips off,
it doesn't trace it
out effectively, OK?
Does everyone see that?
So, it's DNA is always
a right handed helix.
You know, so this beautiful
structure of DNA is one
that was solved by
x-ray crystal structure.
Before then, there were
a large number of wrong,
incorrect predictions about DNA
structure, including by people
who I, you know, think the
world of, I think are, you know,
absolute heroes in science.
For example, the
great Linus Pauling
who proposed a triple
helix of DNA
where the phosphodiester
backbones would be in the center
of the molecule and the bases
would be out on the outside.
This kind-- this is somewhat--
this is intellectually
attractive if you don't think
about the fact that
you have two parents.
But furthermore it's attractive
because at least the base pairs
would be out here in space
where they can interact
with transcription factors.
We now know of course
that that's not correct.
Instead, we'll take
a look in a moment
at where the transcription
factors interact.
Before we do, let's zoom
out a little bit, OK?
So, DNA in the cell is
concentrated in two regions,
a nucleosome in the
prokaryotic cell, so it's kind
of concentrated in the very
center of an E. coli cell.
In a eukaryotic cell of
course, DNA is found exclusive--
is found in the nucleus
and also the mitochondria
but let's just focus on DNA
that's in the nucleus for today.
The bases themselves
are connected together
to form oligonucleotides
through this phosphate,
this phosphodiester
functionalities, OK?
So this is called a
phosphodiester functionality.
The DNA also has a
directionality associated
with it, OK.
So there are-- if
we look closely
at this deoxyribose base,
there is a 5 prime end,
there's a 5 prime
hydroxy over here
and a 3 prime hydroxy over here.
And so, the convention
is to always write DNA
in the direction from
5 prime to 3 prime.
In the same way that we read
English going left to right,
DNA is always read out
5 prime to 3 prime.
This is a really
important convention, OK.
Everyone on the planet follows
this convention and I'm going
to hold you to it as well, OK,
because if you read the DNA
in the opposite direction
you get a different--
or different word
coming out, OK.
It spells something else
that might not be this--
it will almost certainly
not be the same thing
and it might actually
be, you know,
might actually cause
a lot of trouble.
So we're always going to
be reading this 5 prime
to 3 prime directionality, so
this sequence here would be read
out as A, C, G and T, OK,
where the structures of A, C,
G and T are shown here, OK.
Don't bother memorizing
this-- sorry.
Don't bother memorizing
the structures of these.
I'll simply give them to
you on the midterm, OK?
So, at a graduate level,
you should know this.
Mariam will need to know this
for her orals exam, but the rest
of you are in luck because I'm
not going to test you on them
at least for this class, OK.
And again, the-- the
directionality matters a lot.
If there is a 5 prime phosphate,
this 5 prime phosphate
is indicated
by a lower case p. Finally,
last bit of nomenclature,
oligonucleotides that are
connected together are often
referred to as oligos and
that's how I'll describe them.
OK, now I realize oligos
is not the most descriptive
nomenclature because it just
simply means an oligomer
or something, but
that's the convention
that we've been operating
under 50 years, OK.
So oligos will refer
to oligonucleotides.
Typically DNA oligonucleotides
compose of deoxynucleic acid.
OK. Now, even though DNA is--
the bases of DNA are called
bases, it turns out they're not
that basic and few
are protonated
at physiological pH. It's--
this is kind of one of
those historical anomalies.
Here's a bunch of pKa's,
for example starting
with triethylamine.
Here is the pKa of the
protonated triethylamine,
the conjugate acid of
triethylamine, pKa of 10.8.
Here is the pKa of cytosine,
thymine, adenine and guanine
and you can see none of
these would be remotely
considered bases.
Whereas this one over here,
triethylamine definitely a base,
OK, as evidenced by the fact
that its conjugate acid
is, you know, 10.8 pKa.
OK. Question so far?
All right.
Now DNA of course is missing
a 2 prime hydroxyl, OK.
So here is RNA, it has a 2
prime hydroxyl over here.
This 2 prime hydroxyl makes
RNA considerably less stable
than DNA.
I didn't point this out--
let me go back to it--
when we talked earlier
about half lives.
Let me just zoom back
to that really fast.
The half life of RNA
is considerably lower
than the half life of DNA, OK.
So, here is the half life
of RNA, 220,000 years,
whereas the half life of DNA
at 220 million years
is much, much greater.
OK, a thousand fold
difference in stability
for the phosphodiester
backbone of the DNA
versus the phosphodiester
backbone of RNA.
This makes sense, OK?
The 2 prime hydroxyl
of RNA sets you
up for hydrolysis using an
intramolecular attack, OK?
So, here's again the
structure of RNA.
Here's the 2 prime hydroxyl.
This 2 prime hydroxyl
can act as a nucleophile
to attack the phosphodiester
backbone
of the RNA setting up cleavage.
Does anyone want to see
the mechanism of that?
OK, all right, let's
take a quick look.
OK, so in this mechanism--
[ Pause ]
Let me just draw
up the structures
and then I'll blank the board.
OK, one second.
OK. So, in this mechanism,
here's our structure of--
[ Pause ]
OK, so here is our
backbone structure of RNA
and I'm just going to draw
this as base over here, OK?
OK, so if there is any
base that's present,
let's just say hydroxide,
this can deprotonate the 2 prime
hydroxyl, giving us an alkoxide.
[ Pause ]
-- adjacent
to the phosphodiester
backbone of the DNA.
This neighboring alkoxide
can now attack the backbone,
the phosphodiester backbone,
giving you a five-membered
ring intermediate.
OK, which I'll show down here.
[ Pause ]
Five-membered ring intermediate
and this intermediate collapse
leading to cleavage of the RNA.
OK. So here is that collapse.
OK. So, we're going to be
making two strands of RNA
that are separated
from each other.
[ Pause ]
OK, so here's one
strand over here
and then here's the
second strand down here.
[ Pause ]
OK. I'm going to just
differentiate this
as base 1 and base 2.
OK. So, notice that the strand
has actually cleaved apart.
You can then hydrolyze this
phosphodiester backbone,
this phosphodiester back
to a phosphomonoester using
another equivalent of hydroxide.
[ Pause ]
And then finally, collapse
of this tetrahedral intermediate
gives us the product.
OK. Questions about
this mechanism?
All right, now notice again, if
DNA lacks this 2 prime hydroxyl
over here, and I just want to
make this totally explicit,
I'm going to label it 2 prime
hydroxyl, 3 prime, 5 prime.
OK, so DNA lacks
the 2 prime hydroxyl
and therefore does not
have an opportunity
for this intramolecular
nucleophilic attack
on the phosphodiester backbone.
So, for this reason, DNA is
a thousand times more stable
than RNA, right, lacking this
intramolecular nucleophile.
Makes sense?
Questions about this?
OK, let's go back.
Turns out that when you look
at the liability of the bases,
we see actually a
different trend.
OK. And actually I think
I'm going to skip that.
OK, moving on.
OK, I'd like you to
learn what I just told.
Don't worry so much
about the base stability.
DNA bases are subject to
important modifications.
These modifications
have dramatic roles
on the phenotype
of organisms, OK?
So, for example, methyl groups
are often transferred to DNA.
I showed structures
of DNA bases.
Again, they're subject
to massive modification
by methyltransferases
and other modifications.
So, for example, here's
5-methylcytosine over here,
4-methylcytosine and
then N6-methyladenine.
These modifications
can dramatically alter
transcription levels.
They can set up the organism
to transcribe some
genes more often, OK.
So, for example,
lacking pigmentation,
the genes that encode
pigmentation are
in my skin cells,
my epidermal cells,
yet they're not transcribed
very often.
And so, it's likely that my
DNA has not been methylated
in those regions.
However, when I go out and
spend a lot of time in the sun,
I'm getting additional
little spots called freckles
which are resulting
from methylation
of those DNA sequences which in
turn then turns on transcription
of the pigmentation and
results in freckles, OK?
So, the environment, the
environment that you're exposed
to can alter these
transcription patterns.
It's one of the ways that
organisms like ourselves respond
to changes in the environment.
It's a very important
way in fact.
And oftentimes this goes
through methylation of DNA.
This DNA methylation is really
as important as sequence
or genomics, and this is
an area called epigenetics.
That's really an area of very
active research that's taking
place in chemical biology.
OK. So, we've looked
at structures
of the bases themselves,
we've looked at structures
of the phosphodiester backbone,
let's start putting
things together to start
to understand the
structure of DNA.
The bases themselves are
slightly U-shaped, OK.
So, here's a base between A
and T, adenine and thymine.
Notice that this base is
composed of two hydrogen bonds.
Here's a base of G and C which
has three hydrogen bonds.
But notice more importantly
that the bases are U-shaped
or equally importantly,
OK, U-shaped here.
The inside of this U
where the R is going
to be towards the ribose,
the deoxyribose ring,
the inside of this U is going
to form the minor groove
which I've showed you
on an earlier slide.
The outside of the curvy
part of this U is going
to form the major groove.
As you have these U's that are
stacked on top of each other
and each one is slightly offset
with each other, this is--
outside is going to result
in a much bigger helix
than the inside over here, OK?
And here's what this looks like.
OK. So, here's a trace of
the phosphodiester backbone
and then I've highlighted just
one Watson-Crick base pair, OK.
And again, notice
that it's U-shaped,
that there's more section
traced out over on this side,
that will be the major groove,
and the inside will
be the minor groove.
Furthermore, the green arrows
define hydrogen bond donation
and acceptance by the base pair.
And notice that there
is a pattern to this,
that there is an
acceptor-acceptor donor, OK.
So, this is a donor
acceptor donor over here.
So, there's actually a
little bit of a pattern
to whether this is a G on this
side and a C on this side or C
and G on the opposite sides.
So, in other words, A and T
are not the same as T and A
because they're going to
present a different pattern
of hydrogen bonds for molecular
recognition where again,
the proteins are going to be--
the transcription factors
are going to be interacting
over here in the major groove
and small molecules
would be interacting
in this minor groove down here.
I should mention that
there's also some protein DNA
interaction in the minor groove.
It tends to be more
minor, however.
OK, let's take a close
look at one example
of a transcription
factor and how it works.
This is the transcription
factor, Fos-Jun,
it consist of a leucine
zipper which is two helices
that interact with the
DNA like chopsticks, OK.
So, these are fitting
neatly in the major groove.
It turns out the major groove
has exactly the right size
to accommodate an alpha
helical protein, OK.
So, this Fos-Jun is
absolutely perfect.
It fits neatly in
the major groove.
Now, these hydrogen bond
donating functionalities are
going to then read out the
sequence of the DNA and look
for a specific sequence
of DNA to interact with,
trying to form complementary
hydrogen bonds,
trying to form complementary
van der Waals interactions
in this sequence, OK?
Let's take a closer look now
at the forces holding
together the DNA double helix.
Earlier, I alluded to the fact
that AT base pairs
form two hydrogen bonds
and GC base pairs form three.
Which one is stronger,
just, you know,
from a crude approximation?
Yeah, three is stronger
than two, right?
OK, so in addition to this, the
DNA structure is held together
by pi stacking between
the bases.
Again, this is a
face-to-face interaction.
Typically not perfectly
face-to-face,
rather it's typically offset.
And that offset needs the
bases to stack not directly
on each other but slightly
twisted from each other,
setting up this helical
structure
that we're now familiar with.
In order for this base pairing
to take place, the base pairing
that I showed on
the previous slide,
you need a particular tautomer
of these aromatic rings, OK.
And the first one that should
strike you as funny is this one
over here, because you can
imagine another resonant
structure that would make
this C aromatic, right?
Notice that the C has--
is non-aromatic in this
tautomer shown here, right.
It only has two pi electrons
rather than the requisite six
that it would need
to be aromatic.
OK, that's almost-- that's
bizarre to begin with, OK?
So, what's going on here is
that there is a preference
for this tautomer
versus this one.
This one is actually
thermodynamically more stable
and the reason for this is that
the carbon-oxygen double bond
over here is quite strong.
I will tell you that I
think any chemist looking
at this could not have
predicted this in advance,
and in fact actually this
tremendously slowed structure
determination of the
original structure
of DNA back in the 1950s.
Watson and Crick were physicists
and weren't as familiar
with the whole notion
of tautomerization
as their chemical
counterparts were racing
to solve the structure of DNA.
And so for them, this did
not look funny whereas
to us I think it does
look funny, right,
because it lacks aromaticity
whereas a structure
on the left is aromatic.
Again, this happens to be
just a little bit more stable
because of the strength of
the carbon-oxygen double bond,
but I don't think anyone
would have predicted that, OK?
I think now we, you know,
with our 21st century guys,
we could predict it,
but going back in time,
I don't think we could have
predicted it so readily.
Similarly, over here, these
amidines are actually going
to be more stable in
the aromatic structure
than an amidine structure.
And in this case, that's
due to the much poor overlap
between a carbon-nitrogen
double bond
than a carbon-oxygen
double bond, OK.
So, all of these lead
to the base pairs
with the hydrogen
bonding preferences
that are shown here, OK?
Whereas for example, this
is a non-aromatic ring
that could aromatic
if it tautomerize,
but it doesn't prefer
to be tautomerized.
Whereas this one over here seems
to prefer to have an amidine
in this structure
because of the strength
of a carbon-nitrogen
double bond, OK?
And here is another
example of that over here.
This one prefers aromatic
because carbon-nitrogen double
bonds are relatively weak.
OK, pretty interesting.
Unnatural bases, however,
could dramatically shift these
preferences for tautomerization,
and a good example of
this is 5-bromouracil, OK.
So, if this compound
here is fed to organisms,
what happens is an unusual
tautomerization preference
where the enol form of bromo-U
is actually more preferred
than it would be if there
was no bromine over here, OK?
So, most of the time, it forms
the regular base pair, however,
some of the time,
it can actually form
the incorrect base pair
because it can actually more
readily access this enol form
of the base, OK?
So, that's due to the electron
withdrawing functionality
of bromine over here, OK?
That's changing this
tautomerization preference.
The consequences of these
are really dramatic.
Because the Watson-Crick
base paring is not followed
as closely, what ends up
happening is the DNA comes
out with all kinds of bizarre
breaks and lesions, OK.
So here are chromosomes
from a normal organism.
I think it's a hamster
in this case.
And then here's chromosomes
from hamsters that were exposed
to bromouracil and you could
see they have all kinds
of bizarre shapes to them,
things are incorrect, OK.
So, this causes cancer
and breakages in DNA
which then eventually lead
to cancer, cancer cells
and tumors in the organism, OK.
All right, so furthermore,
it turns out that
we can test this--
the importance of the strengths
of these hydrogen bonds
by synthesizing unnatural bases.
So, this is one of the great
things about chemical biology.
If you have this hypothesis
that something is important,
then you could test
that hypothesis
by synthesizing compounds
which are say missing
that key functionality.
So, from Watson and
Crick, we expect to find
that hydrogen bonds are holding
together the structure of DNA
and chemists went out
and synthesized variants
of DNA bases that were lacking
that ability to hydrogen
bond, OK?
Structures of these
are shown here, OK?
So, for example, this compound
here is simply a pyrene in place
of a base and it actually
prefers to base pair
with a missing base
over here, OK?
So, these guys over here,
no hydrogen bonding,
no hydrogen bonding over here
and yet these actually prefer
to pair with each other, OK.
So, you can actually have
completely unnatural bases
missing hydrogen bonds that is--
are yet able to form base pairs
with each other preferentially.
What this tells us is
that there's more going
on in DNA structure than
simply hydrogen bonding.
Hydrogen bonding is a nice
simplifying assumption
for our biochemical friends
or molecular biology friends,
but in actuality,
the pi stacking
of DNA is a driving interaction,
the edge to edge interactions
of aromatic functionalities are
also driving these interactions
between the strands of DNA.
And so, while we can do quite
a bit with hydrogen bonding,
there's quite a bit more
that's left to be explored.
OK, last thought, I've
been showing you--
or it's not last thought,
I've been showing you--
Oh, before I get to that,
here's-- here for example is--
this illustration here
emphasizes the importance
of pi stacking in here, OK?
So, one thing is that bigger
bases tend to pi stack better,
for example, the
guanine base tends
to pi stack better
than say cytosine.
All right, in addition,
I've been showing you
Watson-Crick base pairing
where it's a canonical
base pair,
G's and C's have
three hydrogen bonds,
A's and T's have only two.
Other kinds of hydrogen bonding
possibilities are not only
possible but have been observed.
These were proposed
by Karst Hoogsteen
and we observe this a
lot in RNA structure.
We don't necessarily see this in
DNA, but we definitely see this
in RNA and they're
going to come up later.
So, I'll just show you
the structures here.
This is an alternative
to the usual AT base pair
and this is an alternative
to the usual CG base pair.
This one being driven
by a protonation event,
protonation of this
nitrogen over here, OK?
So, this is actually--
these are sort of edge
to edge interactions
rather than the sort
of neat more typical
Watson-Crick base pair.
OK. Any questions about
the structure of DNA?
Anything, whatsoever?
I want to change gears
then and start talking
about how small molecules
interact with DNA.
The first mode that small
molecules can interact
with DNA is to actually slip
into this pi stack of DNA.
So, aromatic compounds can slide
into the pi stack of the DNA
and we're going to
see the consequences
of this can be quite
destructive.
Let's take a look at some
examples, this is a class
of molecules called
intercalators,
meaning that they intercalate
into the pi stack of the DNA,
they get integrated
into the DNA structure.
So, in order to fit
into this pi stack,
these molecules must
be also hydrophobic
and also aromatic, right?
They will form competing
pi-pi stacking interactions
with the DNA and so they
must also be aromatic.
Note too that in order to force
the way into the pi stack,
these molecules force the DNA's
double helix to slightly unwind
to accommodate the
DNA intercalator.
Here are some examples of this.
These are examples
of intercalators.
Notice that they are
all aromatic compounds.
They're all flat and aromatic
to slide into the pi stack.
Many of these molecules
also have positive charge.
Positive charge is
useful, right,
because DNA with the
phosphodiester backbone
of the DNA is negatively
charged.
This gives the molecule a way
to be attracted in the DNA
through a long range
charged-charged interaction,
right?
So these molecules are
going to seek out DNA
like a homing missile.
And once they slide
into the pi stack,
the consequences
can be pretty bad
or actually fairly useful, OK?
Let me show you an example
of a useful intercalation
over here on the right.
This is actually an agarose
gel which is an important way
that chemical biology
laboratories separate
out DNA structures.
Different DNA sequences can
be separated out on the basis
of their size using
these agarose gels.
I'll show you what that looks
like in a couple
of slides from now.
To visualize the DNA, however,
this molecule over here,
ethidium bromide is
incorporated to the gel
and it gets concentrated
into the DNA
by an intercalation interaction.
So, it slips into the
pi stack of the DNA
and it's a fluorescent molecule,
many aromatic compounds
are fluorescent.
We've talked about
fluorescence before.
And so, you can actually
shine UV light on the gel
and wherever you see
this-- these pinkish bands,
that's where the DNA is present.
And so you can actually take a
razorblade for example and cut
out the DNA of a
particular size.
Here's a couple of
more DNA intercalators.
Here's one that's
designed to intercalate
and then have a little
linker and then intercalate
down below the compound.
Here's what it looks
like structurally,
so there's intercalator, linker,
intercalator up here
for example.
I think that-- this
it right over here.
These are also compounds that
are used to treat cancer.
So, dynemicin, adriamycin are
used as anticancer compounds
or some of the first rounds
of anticancer compounds
that are used as
chemotherapeutics.
And we'll talk more
about their mechanism
of action later in the class.
We're not quite there yet.
OK. Let's stop here.
When we come back next
time, we'll be talking more
about the structure of DNA.
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