Mechanisms of Action of All Major Antibiotic Classes

1. Cell Wall Synthesis Inhibitors

A. Beta-Lactams (Penicillins, Cephalosporins, Carbapenems, Monobactams)

• MOA: Bind covalently to penicillin-binding proteins (PBPs), which are transpeptidase enzymes responsible for cross-linking the glycan strands of peptidoglycan. By blocking these enzymes, beta-lactams prevent cell wall cross-linking, causing osmotic instability and bactericidal cell lysis.
• The beta-lactam ring is the pharmacophore - beta-lactamases cleave this ring and destroy activity.
• Examples: penicillin, amoxicillin, cephalexin, ceftriaxone, meropenem, imipenem, aztreonam

Harrison's Principles of Internal Medicine 22E - beta-lactams "inhibit bacterial cell-wall synthesis by binding to cell-wall transpeptidases, cross-linking enzymes that are also called penicillin-binding proteins (PBPs); PBPs are targets that are unique to bacteria and have no mammalian counterpart."

B. Glycopeptides (Vancomycin, Teicoplanin, Dalbavancin, Oritavancin)

• MOA: Bind to the D-Ala-D-Ala terminus of the lipid-linked peptidoglycan precursor (Lipid II). This physically blocks the transglycosylase reaction (polymerization of glycan strands) and also prevents transpeptidation - acting at a step upstream of beta-lactams.
• Because they work on the cell wall from outside the membrane (no need to cross), they are active against many beta-lactam-resistant organisms.
• Only effective against Gram-positive bacteria (Gram-negatives' outer membrane blocks entry).
• Examples: vancomycin, teicoplanin, telavancin

Goodman & Gilman's - "Vancomycin and other glycopeptides inhibit the polymerization or transglycosylase reaction (A) by binding to the d-alanyl-d-alanine terminus of the cell wall precursor unit attached to its lipid carrier."

C. Bacitracin

• MOA: Cyclic polypeptide that binds C55-undecaprenyl pyrophosphate (the lipid carrier that transports peptidoglycan precursors across the membrane), preventing its dephosphorylation and recycling. This halts delivery of new peptidoglycan building blocks to the cell wall.
• Topical use only (nephrotoxic if systemic).

D. Fosfomycin

• MOA: Inhibits MurA (UDP-N-acetylglucosamine enolpyruvyl transferase), the first committed step in peptidoglycan biosynthesis. It is a phosphoenolpyruvate analog that covalently alkylates a cysteine residue in MurA.

2. Protein Synthesis Inhibitors

A. Aminoglycosides - 30S Inhibitors (Irreversible, Bactericidal)

• MOA: Bind irreversibly to the 30S ribosomal subunit (specifically the 16S rRNA of the 30S subunit). This causes:
  1. Misreading of mRNA - incorrect amino acids are inserted (codon misreading)
  2. Inhibition of translocation - ribosome cannot move along mRNA
  3. Premature termination or production of aberrant proteins that insert into the cell membrane, increasing permeability and enhancing drug uptake
• Bactericidal (unique among protein synthesis inhibitors).
• Require oxygen for uptake - inactive against strict anaerobes.
• Examples: gentamicin, tobramycin, amikacin, streptomycin, neomycin

B. Tetracyclines - 30S Inhibitors (Bacteriostatic)

• MOA: Bind reversibly to the 30S ribosomal subunit (16S rRNA). They block the aminoacyl-tRNA from binding to the ribosomal acceptor (A) site, preventing addition of new amino acids to the growing peptide chain.
• Broad-spectrum: active against Gram-positives, Gram-negatives, atypicals, spirochetes.
• Examples: doxycycline, minocycline, tetracycline, tigecycline (a glycylcycline - overcomes efflux resistance)

C. Macrolides - 50S Inhibitors (Bacteriostatic)

• MOA: Bind reversibly to the 23S rRNA of the 50S subunit (specifically the peptide exit tunnel). They block translocation - the movement of peptidyl-tRNA from the A site to the P site - effectively stalling the ribosome and causing premature dissociation of peptidyl-tRNA.
• Examples: erythromycin, clarithromycin, azithromycin

Miller's Review of Orthopaedics - clindamycin and macrolides "inhibit the dissociation of peptidyl-transfer RNA from ribosomes during translocation (the mechanism is through binding to 50S-ribosomal subunit)."

D. Clindamycin - 50S Inhibitor (Bacteriostatic)

• MOA: Binds the 50S subunit (same 23S rRNA site as macrolides - the peptidyl transferase center). Inhibits peptidyl transferase activity and translocation, blocking elongation.
• Active against anaerobes and Gram-positive cocci; covers MRSA (soft tissue infections).

E. Chloramphenicol - 50S Inhibitor (Bacteriostatic)

• MOA: Binds the 50S subunit at the peptidyl transferase center (23S rRNA). Inhibits peptide bond formation by blocking the aminoacyl-tRNA from positioning correctly in the A site.
• Rarely used systemically due to bone marrow toxicity (aplastic anemia - idiosyncratic).

F. Linezolid (and Tedizolid) - Oxazolidinones - 50S Inhibitor (Bacteriostatic)

• MOA: Unique mechanism - binds to the 23S rRNA of the 50S subunit at the P site, preventing formation of the 70S initiation complex (blocks assembly of the ribosome before translation begins). Specifically prevents the fMet-tRNA from binding to the P site.
• Active against MRSA, VRE, and drug-resistant Gram-positive organisms.
• No cross-resistance with other protein synthesis inhibitors due to unique binding site.

G. Streptogramins (Quinupristin/Dalfopristin)

• MOA: Act synergistically at the 50S subunit.
  • Dalfopristin (Group A) blocks peptidyl transferase
  • Quinupristin (Group B) inhibits translocation and causes premature release of peptide chain
  • Together they are bactericidal, unlike each component alone (which is bacteriostatic)

3. DNA/RNA Synthesis Inhibitors

A. Fluoroquinolones (Bactericidal)

• MOA: Inhibit bacterial type II topoisomerases - specifically DNA gyrase (topoisomerase II) and topoisomerase IV.
  • DNA gyrase introduces negative supercoils ahead of the replication fork (relieves torsional stress) - primary target in Gram-negative bacteria.
  • Topoisomerase IV decatenates daughter chromosomes after replication - primary target in Gram-positive bacteria.
  • Fluoroquinolones stabilize the enzyme-DNA cleavage complex, trapping double-strand breaks and causing lethal DNA fragmentation.
• Examples: ciprofloxacin, levofloxacin, moxifloxacin, ofloxacin, delafloxacin

Katzung's Basic and Clinical Pharmacology 16E - ciprofloxacin "inhibits DNA replication by binding to DNA gyrase and topoisomerase IV."

B. Rifamycins (Rifampin/Rifampicin) - Bactericidal

• MOA: Bind the beta subunit of bacterial DNA-dependent RNA polymerase (rpoB), blocking the initiation of RNA synthesis. They form a stable drug-enzyme complex that physically obstructs the elongation of the nascent RNA chain.
• Used for TB (Mycobacterium tuberculosis), leprosy, and Staphylococcus.
• Induces CYP450 enzymes (major drug interactions).
• Resistance develops rapidly if used alone (single-step mutation in rpoB).

C. Metronidazole (Bactericidal)

• MOA: A prodrug selectively activated in anaerobic/microaerophilic environments. Ferredoxin (in anaerobes) donates electrons to reduce the nitro group of metronidazole, generating toxic free radical intermediates (nitro radical anion). These radicals cause DNA strand breaks and disrupt the DNA helix, leading to cell death.
• Selective for anaerobes (aerobes cannot reduce the drug).
• Also active against protozoa (Giardia, Trichomonas, Entamoeba).

D. Nitrofurantoin

• MOA: Also a prodrug activated by bacterial reductases (nitroreductases). Reactive intermediates are generated that damage multiple macromolecules - DNA, ribosomes, and cell wall proteins simultaneously.
• Concentrated in urine; used only for uncomplicated UTIs.

E. Trimethoprim / Sulfonamides (Co-trimoxazole) - Bacteriostatic

• MOA: Block sequential steps in folate synthesis (a pathway bacteria must synthesize de novo; humans obtain folate from diet):
  • Sulfonamides: Structurally mimic PABA (para-aminobenzoic acid), competitively inhibiting dihydropteroate synthase - the enzyme that incorporates PABA into dihydrofolate.
  • Trimethoprim: Competitively inhibits dihydrofolate reductase (DHFR), preventing conversion of dihydrofolate (DHF) to tetrahydrofolate (THF).
  • Combined (TMP-SMX), they produce sequential blockade of folate synthesis, giving synergistic bactericidal effect.
• Without THF, bacteria cannot synthesize purines and thymidylate - DNA synthesis halts.

4. Cell Membrane Disruptors (Bactericidal)

A. Polymyxins (Polymyxin B, Colistin/Polymyxin E)

• MOA: Cyclic lipopeptides that act like detergents. They bind to the lipopolysaccharide (LPS) and phospholipids of the outer membrane of Gram-negative bacteria, displacing calcium and magnesium ions. This disrupts membrane integrity, causing increased permeability, leakage of cellular contents, and rapid cell death.
• Only active against Gram-negative bacteria (including multidrug-resistant Pseudomonas and Acinetobacter).
• Reserved for last resort due to nephrotoxicity.

B. Daptomycin

• MOA: Cyclic lipopeptide that inserts its lipid tail into the cytoplasmic membrane of Gram-positive bacteria in a calcium-dependent manner. It forms oligomeric channels/pores that disrupt membrane potential, causing rapid efflux of potassium ions and membrane depolarization, leading to cell death.
• Active only against Gram-positive organisms including MRSA and VRE.
• Inactivated by pulmonary surfactant - cannot be used for pneumonia.

5. Mycobacterium-Specific Antibiotics

Summary Table