Here we have a penicillin-binding protein
shown as a green ovoid there with PBP, the
abbreviation for the penicillin-binding protein.
I’ve also shown that it’s got CH2OH. This
correlates to the serine amino acid which
is at its active site.
Now, if you recall, we said that the terminus
which is activated of the polypeptide contained
two alanine groups: D-Alanine-D-Alanine. I’ve
shown those here. I’ve also shown them attached
to the peptide chain, but I haven’t shown
the rest of the sugar because there really
isn’t enough space. The way in which penicillin-binding
protein works is it attacks the carbonyl-carbon
of that amide and it forms an ester group
with the second amine... second alanine on
that polypeptide and kicks off the alanine,
as you can see here on the right hand side.
Look at what we’ve done. We’ve converted
an amide into an ester. And do you remember
how we were talking back in Module III, how
that’s a very difficult to achieve from
a synthetic perspective? This is the advantage
in this case of the enzymatic activity for
this biological process because it is able
to achieve this. It converts it into an ester
via a covalent bond formed and the terminal,
the alanine, diffuses away.
What then happens is that the neighbouring
pentaglycyl unit attached to the other polysaccharide
chain then comes in and attacks that carbonyl-carbon
via an addition-elimination reaction. What
then happens is the formation of an amide
and the diffusing away of the penicillin-binding
protein as a reactivated enzyme to begin again.
Pay particular attention to the bottom right
hand corner where I show what we’ve done
is we’ve linked a polysaccharide to one
peptide chain to, then, the peptide chain
with the other polysaccharide. We have cross-linked
the polysaccharide chains using these polypeptide
bridges. Once the enzyme is restored, it can
carry on and do its thing.
Penicillins, though, work to inhibit this
process. And they’re thought to resemble
closely the geometry of an acylated D-Alanine-D-Alanine
dipeptide. As you can see here on the right
hand side, penicillin or the general structure
for it, is shown to the middle on the right
hand side and at the bottom right hand corner,
an acylated D-Alanine-D-Alanine is shown.
As you can see, the stereochemical orientation
is remarkably similar and this forms a key
part of it as a pharmacophore for inhibiting
the penicillin-binding protein.
Transamidase or penicillin-binding protein
mistakes it for its normal substrate and as
we’ll see in a second, this results in the
irreversible inhibition of the penicillin-binding
protein and prevents the crosslinking of the
peptidoglycan. Because of the ring system,
the hydrolysis does not result in penicillin
breaking into two units like a D-Alanine-D-Alanine
dipeptide would. And the heterocyclic ring,
this thiazolidine ring, is thought to be a
steric barrier to the approach of the pentaglycyl
unit. As a consequence of this, the enzyme
is irreversibly inhibited and cross-links
cannot form. Resulting in this, is that the
cell wall becomes structurally weak, leading
to cell lysis and death. Because, bear in
mind, the bacteria is under constant attack
from the host’s own immune system.
So, let’s have a look at that in a bit more
detail, shall we?
So, here we’ve got the diagram I showed
you before. We’ve got the D-Alanine-D-Alanine
cell-wall fragment. We’ve got the serine
hydroxyl and the active site of the penicillin-binding
protein. What happens in this scenario is
that we get attack by the serine hydroxyl
group, as we showed before, which cleaves
in between the D-Alanine-D-Alanine dipeptide,
converting it into an ester and the terminal
D-Alanine diffuses away.
Under normal circumstances, a pentaglycyl
unit attached to the neighbouring polysaccharide
and peptide chain would enter the active site
and attacks the enzyme peptidoglycan complex,
thus resulting in amide-bond formation and
regeneration of our penicillin-binding protein.
However, in the presence of penicillin what
happens is this. The serine hydroxyl, mistaking
the penicillin for its natural substrate,
attacks the beta-lactam group at the carbonyl-carbon,
thus opening up that carbon-oxygen double
bond and breaking open the nitrogen-carbon
double bond. As you can see here in this scenario
in the bottom left hand corner, we have now
generated a penicillin-binding protein which
has been esterified by the penicillin, forming
an ester bond here. Now, of course, you might
think well it’s an ester, surely that could
hydrolyse. But, that’s where the thiazolidine
ring comes in, ostensibly blocking the active
site and preventing other molecules from entering.
This enzyme is now inactive and has to be
broken down and reformed by the bacteria.
So, that’s brilliant. But, there are problems.