Peptidoglycan – Beta Lactam Antibiotics

00:01 So, let’s talk about how it actually works. It works by targeting peptidoglycan. Peptidoglycan is a mixed heteropolymer of hexose sugars and amino acids which effectively surrounds a bacterium like a net. The sugars are made up of N-acetylglucosamine, which is abbreviated NAG, and N-acetylmuramic acid, abbreviated NAM. And they’re linked alternately in a chain or polysaccharide. Attached to the NAM or N-acetylmuramic acid are short peptides.

00:40 So, let’s have a quick look of what this means from a physiological perspective to some extent.

00:47 Here we’ve got an example of two types of bacteria. On the left hand side, we have the Gram-positive and on the right hand side, we have the Gram-negative. And these are broad classifications of bacteria that are encountered.

01:01 Gram-positive bacteria, for example, such as Staphylococcus Aureus, and Gram-negative bacteria, for example, Pseudomonas Aeruginosa. So, let’s have a look at the differences between the two.

01:15 If we zoom into the cell wall in both types of bacteria, we see that the cell wall in Gram-positive bacteria has a very thick layer of peptidoglycan (shown in purple) that protects the cell membrane.

01:28 If we look on the right-hand side, we see that the cell wall in Gram-negative bacteria has a lipopolysaccharide, which is over the top of the outer membrane and thin peptidoglycan layer.

01:40 This does or has presented a problem with the use of penicillins in the treatment of Gram-negative bacterial infections. But, we’ll come onto that a little later on.

01:48 So, being aware of this and being shown this in cartoon form, what I’d like to now do is drill down into the details of the structure of the peptidoglycan. And this brings me to the next slide.

02:00 Here we have balls representing the N-acetylmuramic and the N-acetylglucosamine shown as NAM and NAG respectively. Here we show that the NAM units are linked together with NAM units on adjacent chains by the short peptide bridges. Now, this is a good cartoon structure which makes you understand how the layers of peptidoglycan are built up around bacteria. But, it does nothing in terms of explaining what’s happening on a molecular level. So, let’s zoom in a little more.

02:35 If we zoom in a little more and we look at our NAG-NAM backbone, which we showed as the alternating balls of our sugar backbone, we can see that the linkers are constituted of a variety of different amino acids shown here: L-Alanine, D-Glutamate, L-Lysine, followed by this pentaglycyl polypeptide which links to, again, D-Alanine, L-Lysine, D-Glutamate and L-Alanine. And it’s this chain here which is the important point of action for our target enzyme.

03:12 What do I mean by that? Irreversible inhibition of the enzymes involved in cross-linking peptidoglycan is the main mode of action for penicillins. Transamidase, which is a penicillin-binding protein cleaves between two D-Alanine-D-Alanine amino acids at the end of the peptide from the N-acetylmuramic acid (NAM). Attack by the free amino end of a pentaglycyl unit of an adjacent peptide thus forms a crosslink between the two linking the chains together.

03:50 So, let’s have a look at a diagram since, sometimes, a mechanism speaks a thousand words.

03:55 What’s happening in this particular case is that the enzyme or penicillin-binding protein activity is cleaving off that terminal D-Alanine and activating the one next to it. What then happens is the pentaglycyl unit from a neighbouring chain creates an amide bond or peptide bond between itself and that penultimate D-Alanine. This results in the cross-linking of the two chains. Now, we have two sugar backbones held together by a polypeptide bridge and it’s this structural integrity that the penicillin-binding protein is responsible for. And it is this part that is responsible... that penicillins are shown to inhibit.

04:45 Okay. So, we’ve actually looked at it from an amino acid perspective and that’s fine.

04:48 But, let’s drill down even further and start looking at it from a proper molecular perspective.