MECHANISMS OF ACTION OF

ANTIMICROBIAL AGENTS

DR.MUSA EZEKIEL

DEPT.OF MEDICINE

SYNOPSIS

• Historical background

• Classification

• Mechanisms of action

• Conclusion

HISTORY

• Antimicrobial agents are substances that are used to inhibit or kill the growth of microorganisms(bacteria,viruses,fungi and protozoa)

• The earliest evidence of successful chemotherapy is from ancient Peru, where the Indians used bark from the cinchona tree to treat malaria.

• Other substances were used in ancient China

HISTORY-2

17th Century:

Treatment of infectious diseases, e.g.

• quinine for malaria;

• emetine for amoebiasis.

HISTORY-320th century:

• Modern chemotherapy has been dated to the work of Paul Ehrlich in Germany,

• Paul Ehrlich formulated the principle of selective toxicity.• The first planned chemotherapy was arsphenamines for

syphilis.1935:• The beginning of Current era of chemotherapy with

discovery of the sulfonamides.1940:• Penicillin that was discovered in 1929, was demonstrated to

be effective Subsequent 25 years after: • Development of streptomycin, tetracycline,

chloramphenicol and others.

HISTORY-4

Recently:

• Biosynthesis- modifications of molecules for development of new antimicrobial agents.

Classification of antimicrobial agents

• According to whether they are bactericidal or bacteriostatic

• By target site

• By chemical structure

ANTIMICROBIAL AGENTS

Selective toxicity• Exhibited by an ideal antimicrobial agent.• Drug is harmful to the parasite but not to the

host.• relative rather than absolute, i.e. drug in

concentration tolerated by the host may damage an infecting microorganism.

• due to a function of drug specific receptor or inhibition of biochemical events essential to the organism.

Biochemical Basis of Antimicrobial Action • Bacterial cells grow and divide, replicating

repeatedly to reach the large numbers present during an infection or on the surfaces of the body.

• To grow and divide, organisms must synthesize or take up many types of biomolecules.

• Antimicrobial agents interfere with specific processes that are essential for growth and/or division .

Biochemical Basis of Antimicrobial Action -3

Antibiotic agents may be either:• bactericidal:- killing the target bacterium• Bacteriostatic:- inhibiting its growth.

Bactericidal agents are more effective, but bacteriostatic agents can be extremely beneficial since they permit the normal defenses of the host to destroy the microorganisms.

MECHANISMS OF ACTION OF ANTIMICROBIAL AGENTS

They are classified as follows:

• inhibitors of bacterial cell walls,

• inhibitors of cytoplasmic membranes,

• inhibitors of nucleic acid synthesis, and

• inhibitors of ribosome function .

• Inhibitors of folate pathway

Inhibition of cell wall synthesis(Bacitracin, Cephalosporins, Cycloserine, Penicillins, Vancomycin)• Bacteria are classified as Gram-positive and

Gram-negative organisms on the basis of staining characteristics.

• Gram-positive bacterial cell walls contain peptidoglycan and teichoic or teichuronic acid, and the bacterium may or may not be surrounded by a protein or polysaccharide envelope.

• Gram-negative bacterial cell walls contain peptidoglycan, lipopolysaccharide, lipoprotein, phospholipid, and protein .

Inhibition of cell wall synthesis-2

• The critical attack site of anti-cell-wall agents is the peptidoglycan layer.

• This layer is essential for the survival of bacteria in hypotonic environments;

• loss or damage of this layer destroys the rigidity of the bacterial cell wall, resulting in death.

Inhibition of cell wall synthesis-3

• The rigid cell wall possessed by most bacteria is lacking in the host cells. This is prime target for agent that exhibit selective toxicity.

• Inhibitors of bacterial cell wall synthesis act on the formation of peptidoglycan layer.

• Bacteria that lack peptidoglycan, such as mycoplasmas, are resistant to these agents.

Inhibition of cell wall synthesis-5

• Penicillins and cephalosporins/cephamycins are widely used to inhibit both Gram-positive and Gram-negative bacilli.

• Monobactams inhibit only aerobic Gram-negative bacilli,

• Clavulanic acid acts as a ß-lactamase inhibitor, and thienamycin inhibits a wide range of aerobic and anaerobic species.

Inhibition of cell wall synthesis-6• Vancomycin interrupts cell wall synthesis by forming

a complex with the C-terminal D-alanine residues of peptidoglycan precursors.

• Complex formation at the outer surface of the cytoplasmic membrane prevents the transfer of the precursors from a lipid carrier to the growing peptidoglycan wall by transglycosidases.

• Biochemical reactions in the cell wall catalyzed by transpeptidases and D,D-carboxypeptidases are also inhibited by vancomycin and other glycopeptide antimicrobials.

• Because of its large size and complex structure, vancomycin does not penetrate the outer membrane of gram-negative organisms.

Antibiotics that Affect the Function of Cytoplasmic Membranes

Bacterial Cytoplasmic Membranes

• Biologic membranes are composed basically of lipid, protein, and lipoprotein.

• The cytoplasmic membrane acts as a diffusion barrier for water, ions, nutrients, and transport systems.

• Most workers now believe that membranes are a lipid matrix with globular proteins randomly distributed to penetrate through the lipid bilayer.

Bacterial Cytoplasmic Membranes-2

• A number of antimicrobial agents can cause disorganization of the membrane. These agents can be divided into cationic, anionic, and neutral agents.

• The best-known compounds are polymyxin B and colistemethate (polymyxin E).

• These high-molecular-weight octapeptides inhibit Gram-negative bacteria that have negatively charged lipids at the surface.

Bacterial Cytoplasmic Membranes-3

• Since the activity of the polymyxins is antagonized by Mg2+ and Ca2+, they probably competitively displace Mg2+ or Ca2+ from the negatively charged phosphate groups on membrane lipids.

• Basically, polymyxins disorganize membrane permeability so that nucleic acids and cations leak out and the cell dies.

• The polymyxins are of virtually no use as systemic agents since they bind to various ligands in body tissues and are potent toxins for the kidney and nervous system.

Antibiotics that Inhibit Nucleic Acid Synthesis

Antimicrobial agents can interfere with nucleic acid synthesis at several different levels.

• inhibit nucleotide synthesis or interconversion;

• prevent DNA from functioning as a proper template; and

• interfere with the polymerases involved in the replication and transcription of DNA.

Inhibition of DNA-Directed RNA Polymerase

• Rifamycins are a class of antibiotics that inhibit DNA-directed RNA polymerase.

• Polypeptide chains in RNA polymerase attach to a factor that confers specificity for the recognition of promoter sites that initiate transcription of the DNA.

• Rifampin binds noncovalently but strongly to a subunit of RNA polymerase and interferes specifically with the initiation process.

• However, it has no effect once polymerization has begun.

Inhibition of DNA Replication

• DNA gyrase and topoisomerase I act in concert to maintain an optimum supercoiling state of DNA in the cell. In this capacity,

• DNA gyrase is essential for relieving torsional strain during replication of circular chromosomes in bacteria.

• The enzyme is a tetrameric protein composed of two A and two B subunits. A transient, covalent bond between the A subunit and DNA occurs during the double strand passage reaction catalyzed by gyrase.

Inhibition of DNA Replication-2• Quinolones such as nalidixic acid, bind to the

cleavage complex composed of DNA and gyrase during this strand passage.

• This interaction of quinolone acts to stabilize the cleavage intermediate which has a detrimental effect on the normal DNA replication process.

• The effects of this inhibition result in the death of the bacterial cell.

• The newer fluoroquinolones such as ciprofloxacin, norfloxacin, and ofloxacin also interact with DNA gyrase and possess a broad spectrum of antimicrobial activity.

Inhibition of DNA Replication-3

• Nalidixic inhibits only aerobic Gram-negative species.

• In ciprofloxicin, the flourine provides Gram-positive activity, the piperazine group increases activity against members of the Enterobacteriaceae, and the piperazine and cylopropyl groups give activity against Pseudomonas species.

Inhibition of DNA Replication-4

• Nitroimidazoles such as metronidazole inhibit anaerobic bacteria and protozoa. The nitro group of the nitrosohydroxyl amino moiety is reduced by an electron transport protein in anaerobic bacteria. The reduced drug causes strand breaks in the DNA. Mammalian cells are unharmed because they lack enzymes to reduce the nitro group of these agents.

• Metronidazole enters an aerobic bacterium via the electron transport protein ferrodoxin,where it is reduced. The drug then binds to DNA, and DNA breakage occurs.

Antimicrobial Inhibitors of Ribosome Function.

• Bacterial ribosomes contain two subunits, the 50S and 30S subunits.

• Anti-ribosomal antibiotics impair ribosomes by binding to either 50S or 30S ribosomal subunits

• Ribosomes are essential for translation of mRNA into proteins

• No translation No protein synthesis• No protein synthesis No growth

Ribosome Home Plate

• Baseball player slides into home

• The ball is fielded by the catcher who makes a CLEan TAG

• The word CLEean lies over the base: these inhibit 50S

• The word TAG lies beneath the base: these inhibit 30S

Antimicrobial Inhibitors of Ribosome Function-2

• Aminoglycosides act by binding to specific ribosomal subunits.

• Aminoglycosides are complex sugars connected in glycosidic linkage .

• They differ both in the molecular nucleus, which can be streptidine or 2-deoxystreptidine, and in the aminohexoses linked to the nucleus.

• Essential to the activity of these agents are free NH, and OH groups by which aminoglycosides bind to specific ribosomal proteins.

Antimicrobial Inhibitors of Ribosome Function-3

• Streptomycin was the first aminoglycoside studied

• It is rarely used clinically today except to treat tuberculosis.

• Its mode of action differs to some extent from that of the other clinically useful aminoglycosides, which are 2-deoxystreptidine derivatives such as gentamicin, tobramycin, and amikacin.

• Streptomycin binds to a specific S12 protein in the 30S ribosomal subunit and causes the ribosome to misread the genetic code.

Antimicrobial Inhibitors of Ribosome Function-4

• Other aminoglycosides bind not only to the S12 protein of the 30S ribosome, but also to some extent to the L6 protein of the 50S ribosome.

• This latter binding is quite important in terms of the resistance of bacteria to aminoglycosides.

• Indeed, the aminoglycoside-type drugs can combine with other binding sites on 30S ribosomes, and they kill bacteria by inducing the formation of aberrant, nonfunctional complexes as well as by causing misreading.

Antimicrobial Inhibitors of Ribosome Function-5

• Other agents that bind to 30S ribosomes are the tetracyclines.

• These agents appear to inhibit the binding of aminoacyl-tRNA into the A site of the bacterial ribosome.

• Tetracycline binding is transient, so these agents are bacteriostatic.

• Nonetheless, they inhibit a wide variety of bacteria, chlamydias, and mycoplasmas and are extremely useful antibiotics.

Antimicrobial Inhibitors of Ribosome Function-6

• Spectinomycin is an aminocylitol antibiotic that is closely related to the aminoglycosides. It binds to a different protein in the ribosome and is bacteriostatic but not bactericidal. It is used to treat penicillin-resistant gonorrhea.

Antimicrobial Inhibitors of Ribosome Function-7

• There are three important classes of drugs that inhibit the 50S ribosomal subunit.

• Chloramphenicol is a bacteriostatic agent that inhibits both Gram-positive and Gram-negative bacteria. It inhibits peptide bond formation by binding to a peptidyltransferase enzyme on the 50S ribosome.

• Macrolides are large lactone ring compounds that bind to 50S ribosomes and appear to impair a peptidyltransferase reaction or translocation, or both.

• The most important macrolide is erythromycin, which inhibits Gram-positive species and a few Gram-negative species such as Haemophilus, Mycoplasma, Chlamydia, and Legionella.

Antimicrobial Inhibitors of Ribosome Function-8

• New molecules such as azithromycin and clarithromycin have greater activity than erythromycin against many of these pathogens.

• Lincosamides, of which the most important is clindamycin, have a similar site of activity

• Both macrolides and lincosamides are generally bacteriostatic. inhibiting only the formation of new peptide chains.

Inhibitors of folate pathway

• Sulfonamides competitively block the conversion of pteridine and p-aminobenzoic acid (PABA) to dihydrofolic acid by the enzyme pteridine synthetase.

• Sulfonamides have a greater affinity than p-aminobenzoic acid for pteridine synthetase.

• Trimethoprim has a tremendous affinity for bacterial dihydrofolate reductase (10,000 to 100,000 times higher than for the mammalian enzyme); when bound to this enzyme, it inhibits the synthesis of tetrahydrofolate.

Antibacterial Agents that Affect Mycobacteria

Isoniazid• is a nicotinamide derivative that inhibits mycobacteria. • precise mode of action is not known, but it affects the

synthesis of lipids, nucleic acids, and the mycolic acid of the cell walls of these species.

Ethambutol • mechanism of action is unknown. • mycostatic, whereas isoniazid is mycocidal. Rifampin and streptomycin, • affect mycobacteria in the same manner that they inhibit

bacteria. Pyrazinamide • a synthetic analog of nicotinamide. It is bactericidal, but its

exact mechanism is unknown

Conclusion

• Adequate knowledge of various antimicrobial classification,mechanisms of action are crucial in optimal patients care and in prevention of resistance.

THANK YOU

FOR

LISTENING