Antibiotics; Inhibitors of Cell Wall Synthesis LECTURE 22: Antibiotics; Inhibitors of Cell Wall Synthesis Microbiology and Virology; 3 Credit hours Atta-ur-Rahman School of Applied Biosciences (ASAB) National University of Sciences and Technology (NUST)

Antibiotics Antibiotics are agents that are "selectively" toxic for bacteria (either killing them [bactericidal] or inhibiting their growth [bacteriostatic]) without harm to the patient. Antibiotics work most efficiently in conjunction with an active immune system to kill infecting bacteria in the host. The minimal inhibitory concentration (MIC) refers to the lowest concentration of an antibiotic that stops visible growth.

Components of Cell Wall

Peptidoglycan Peptidoglycan consists of polysaccharide backbone consisting of N-acetyl muramic acid and N-acetyl glucosamine with peptide side chains containing D- and L- amino acids and in some instances diaminopimelic acid. The side chains are cross-linked by peptide bridges by Penicillin binding protein (PBP).

Synthesis of Cell Wall The precursor subunit (muramyl pentapeptide attached to uridine diphosphate, UDP) is synthesized in the cytoplasm and passed to the cell membrane. The subunit is moved enzymatically to a lipid carrier (undecaprenol) and built into a completed subunit (disaccharide pentapeptide with attached bridge peptide). The completed subunits are then exported to the cell wall. After release of the monomer the undecaprenol is recirculated in the cell membrane and used again.

Synthesis of Cell Wall The glycan backbones of the existing cell wall is enzymatically broken (by autolysins) to allow insertion of the newly synthesized subunit. Cross-linking of the peptide side-chain of the inserted subunit to the existing chain then occurs enzymatically (penicillin binding proteins).

Synthesis of Lipopolysaccharide Lipid A is assembled in the cell membrane and the core sugars attached sequentially. O-antigen subunits are independently synthesized (on a lipid carrier as in peptidoglycan synthesis). The fully synthesized O-antigen is then attached to the lipid A-core (generating lipopolysaccharide) in the cell membrane before insertion into the outer membrane.

Antibiotics; Mechanism

1- Cycloserine; Inhibition of peptidoglycan synthesis The terminal two amino acids of a peptide side chain of peptidoglycan are unusual amino acids (D-alanine as opposed to its isomer L-alanine). The antibiotic cycloserine is an analog of D-alanine and interferes with enzymatic conversion of L-alanine to D-alanine in the cytoplasm. Thus, subsequent synthesis of peptidoglycan cannot occur.

Cycloserine; Antibiotic

2- Vancomycin; Inhibition of peptidoglycan synthesis Cross-linking of the peptide occurs in the cell wall. During this process D-alanine is enzymatically excised from the end of a pre-existing peptide side chain allowing it to be cross-linked (by a new peptide bond) to the recently synthesized peptidoglycan subunit. Vancomycin  binds to D-alanine-D-alanine thus sterically inhibits transpeptidation (cross-linking).

Vancomycin

3- β-Lactam; Inhibition of peptidoglycan synthesis The beta lactam antibiotics include penicillins (e.g. ampicillin), cephalosporins and monobactams and carbapenems. They bind to and inhibit enzymes (penicillin binding proteins) involved in the transpeptidation (cross-linking) of peptidoglycan. These antibiotics have in common the four membered lactam ring. Attached to the lactam, penicillins have an additional five membered ring and cephalosporins a six membered ring. Monobactams consist of the lactam ring alone and display antibiotic activity.

β-Lactam Antibiotics Penicillin nucleus Benzyl penicillin Monobactam nucleus Ampicillin Aztreonm Carbapenem nucleus Imipenem

4- Bacitracin; Inhibition of peptidoglycan synthesis The peptidoglycan subunit is passed across the cytoplasmic membrane attached to undecaprenol diphosphate. After the nascent peptidoglycan monomer leaves the carrier on reaching the cell wall, the undecaprenol diphosphate is dephosphorylated to its monophosphate form. Bacitracin inhibits the dephosphorylation reaction and in the absence of monophosphorylated carrier peptidoglycan subunit synthesis stops. 

Bacitracin

5- Polymyxin B ; Inhibition of peptidoglycan synthesis It is derived from the bacterium Bacillus polymyxa. Polymyxin B binds to the lipid A portion of lipopolysaccharide and also to phospholipids. However, it binds preferentially to lipid A. This disrupts the outer membrane of Gram negative bacteria. Since the cell membrane is not exposed in Gram positive bacteria polymyxin has little activity against them. This drug is toxic to human cells, since it can also lyze eukaryotic membranes; this explains its limited clinical use.