Phosphodiesters: Phosphorus in Nucleic Acids

Uploaded by lamechivanes on 13.12.2010


There is a world of difference in the reactivity
between phosphomonoesters that we talked about
in the last webcast
and the phosphodiester functional group
that is the topic of this webcast.
The phosphodiester functional group
links together two organic consituents
through oxygen and phosphorus atoms.
The two remaining oxygens are going to normally carry
a negative charge.
The reason is- is because if that oxygen were protonated,
it has a very low pKa, meaning it’s a very strong acid.
It’s actually stronger than phosphomonoesters
or phosphoric acid itself.
We’ve encountered phosphodiesters
when we talked about nucleic acids.
There you can see the structure of DNA.
The two organic constituents
are the two different ribose rings
that are linked together
through that phosphodiester linkage.
We talked about DNA and RNA as being polyanionic,
and the reason is is because that proton is never
usually bound to that oxygen being so acidic.
It’s almost always lost
and forms the polyanionic backbone of nucleic acids.
Another example of where phosphodiester linkage is-
shows up in biological chemistry is in lipid bilayers.
For example, phosphatidylcholine
which has the ah, two different organic constituents
shown here.
One is attached to a very long chain fatty acids
that are hydrophobic, and the other side is very polar.
It contains this trialkylammonium group,
so that, overall, that negative charge on the phosphodiester
is offset by the positive charge on the trialkyl group,
meaning that it’s a zwitterionic entity.
This part of the molecule,
unlike the other portion of the molecule over here,
is very, very hydrophilic
to, compared to this very hydrophobic entity,
and that’s how the amphiphilic membrane
forms its bilayer structure.
The hydrophobic groups back up back to back
from two different phosphotidylcholine molecules,
forming the bi- lipid bilayer structure.
While I briefly mentioned that there’s a world of difference
in reactivity between the mono and di phosphoesters,
and it’s summarized for you here.
The monoesters are really quite easily hydrolyzed;
whereas, the phosphodiesters
are quite resistant to hydrolysis.
And this turns out to be a very important thing
for life itself because the phosphodiester linkage
is necessary in order to carry information
from one generation of a cell to the next.
If, since the sequence of DNA
is what carries that information
if this linkage underwent hydrolysis,
the fidelity of that information would be lost.
It’s been estimated
that the hydrolysis of the phosphodiesters
is not only stable on the cell lifetime,
but it’s actually stable on much longer life,
much longer scales.
For example, prehistoric time scales is
estimated to be the half-life
of the hydrolysis of the phosphodiester group.
Under physiological conditions and 37º,
about 80 million years is the hydrolysis half-life
for the phosphodiester group.
Let’s look at the nonenzymatic hydrolysis mechanisms,
first by looking at the mechanism
that we talked about last time.
That SN2 process in which the phosphomonoester
undergoes hydrolysis, and the key thing to remember
is that that proton that was available
was able to serve as an intramolecular general acid
so that the leaving group could leave in its neutral form.
It was a one-step process and so all we talked about
was the transition state structure.
In the case of phosphodiesters,
there is no proton to serve as a general acid,
and so that leaving groups leaves as this alkoxide anion.
A consequence of that
is that a different reaction pathway ensues.
This pathway, the SN2 like pathway,
is no longer available.
Instead, the pathway follows an addition-elimination process
much like what we’ve encountered for typical
carbon-oxygen double bond carbonyl kinds of chemistry.
So instead, there is this
trigonal bipyramidal intermediate
which is a fully formed intermediate where all bonds
are fully formed unlike the transition state structure
where bonds are being formed and being made,
and there isn’t, ah, in the case of the phosphomonoester,
any intermediate at all.
So the key difference just to summarize,
emphasize that point is the nature of the leaving group,
and on the next slide I summarized for you
the similarities and the differences.
Now the similarities,
evidence indicates that the transition states
– so in the case of the phosphomonoester
there’ll only be a single transition state.
In the case of the phosphodiester,
there’ll be two transition states,
but the evidence indicates that all of the transition states
involve this oxydianion transition state structure,
meaning that there’s two negative charges
in the transition state.
And so the negative charges are present
in both of the transition states that are found
for the phosphodiester hydrolysis mechanism.
The outgoing group here
is picking up the negative charge that-
the second negative charge that existed, ah, from that,
ah, trigonal bipyramidal intermediate.
As far as – so those are the similarities.
As far as the difference goes,
the key difference that I really want to emphasize
is the one that’s shown here,
and that is the difference in pKa.
Ah, keep the main thing the main thing.
Well, the main thing is the pKa difference
between the outgoing leaving groups.
We have a neutral alcohol
in the case of the phosphomonoester.
Whereas, the leaving group for phosphodiesters
must be the alkoxide.
Above all else, above everything else,
this is the main point that you should keep in mind.
It is the reason DNA is stable as opposed to phosphate,
phosphomonoesters which are, are not stable.
These other effects are small.
There’s ah delocalization over three oxygen.
Ah, the negative charge is delocalized over three oxygens
in the case of the phosphomonoester.
That simply means that this oxygen is actually picking up
some of the negative charge as this proton departs,
and so we can see that the negative charge is spread over
those three oxygen atoms.
In the case here, that, ah, oxygen is essentially tied up,
ah, with its organic constituent,
so it’s not going to pick up any negative charge.
And then as far as the pKa difference goes,
these are small effects.
The big effect is that leaving group effect
that’s shown here.