Question

Part II – Resistance

Among the first antibiotics used on a large scale was penicillin, which was discovered in 1929 by Alexander Fleming.  It was finally isolated and synthesized in large quantities in 1943. Penicillin works by interfering with the bacterial cell wall synthesis. Without a cell wall, the bacterial cells cannot maintain their shape in changing osmotic conditions. This puts significant selective pressure on the microbes to evolve, as they cannot survive the osmotic stress. Any microbe that can resist these drugs

will survive and reproduce more, making the population of microbes antibiotic resistant.

The specific mechanism of penicillin is the prevention of cell wall

synthesis by the ß-lactam ring of the antibiotic (Figure 3), which binds and

inhibits an enzyme required by the bacterium in this process.

The enzyme is called penicillin-binding protein (PBP), even though it is an

Enzyme involved in cell wall synthesis. Normally enzymes have names that

indicate what they do and end in the suffix -ase, like lactase, the enzyme

that breaks down lactose. Figure 4 is a representation of PBP and its active

site.

Bacteria cell walls are layered structures, where each layer is made of peptidoglycan, a sugar and protein polymer. Each layer is cross-linked to the next, strengthening the wall and allowing the cell to resist osmotic pressure. The way the enzyme PBP works is to form those cross-bridges by joining strings of amino acids together in the active site, which is a groove in the protein (Figure 5).

The PBP takes amino acid residues attached to peptidoglycan layers and forms

Brides between the within the active site groove. This cross-linking, or cross-

bridging, stabilizes and strengthens the cell wall. ß-lactam antibiotics interfere

with the PBP enzyme by binding to the active site, blocking the site from the

amino acids (Figure 6).

There are over 80 natural and semi-synthetic forms of ß-lactam antibiotics, including cephalothin and methicillin. Vancomycin also interferes with cell wall synthesis, but its mechanism of action is to bind directly to the cell wall components (Figures 7 and 8).

The first MRSA case was discovered in 1961 in a British hospital, and was the result of a mutation in the enzyme normally inhibited by the ß-lactam ring of methicillin. The site where the antibiotic would bind no longer allowed access to the ring, so the enzyme continued to function normally. The microbe acquired a new gene that, when made into protein, was a different version of PBP, one that couldn’t be inhibited by penicillin.

Question

4. What is the difference in how ß-lactam antibiotics and vancomycin work?

Answer 1

Both  ß-lactam and Vancomycin acts by inhibiting cell wall synthesis by the mechanism is different.

Bacterial cell wall is composed of peptidoglycan which has glycan chain crosslinked with peptide chain. The glycan chain is composed of alternating amino sugars, NAM (N-acetyl Muramic acid) and NAG (N-acetyl Glucosamine). A pentapeptide linked to NAM, has pentaglycine attached to it. This pentaglycine is crosslinked with pentapeptide of adjacent strand. Transpeptidase (Penicillin Binding Protein) causes crosslinking of pentaglycine residue of one strand and fourth amino acid (D-alanine) of adjacent strand, by clevage of the terminal D-alanine (5th amino acid). This makes the cell wall rigid and stable.

Mechanism of  ß-lactam :-

ß-lactam are structural analogues of D-alanine, inhibits Transpeptidase.  ß-lactam block the clevage of terminal D-alanine and transpeptidation. This leads to Inhibition of crosslinking of peptidoglycan and cell wall Synthesis.

Mechanism of vancomycin :-

Vancomycin binds tightly to the terminal D-Ala-D-Ala sequence and prevents its release from the bactoprenol lipid carrier so that the assembly of the units at the cell membrane and crosslinking cannot take place.

The peptidoglycan units that are synthesized within the bacteria cell are transported across the cell membrane by attachment with a bactoprenol lipid carrier for assembly into strand.

Since vancomycin binds to the terminal dipeptide it will prevent its release and further transpeptidation. Thus inhibits further elongation and crosslinking of the peptidoglycan matrix.

This is the difference that how  ß-lactam and vancomycin works.

The diagram given below will depicts the site of action of  ß-lactam and vancomycin :-