Antimicrobials 1:
Origins and modes of action
Dr Fiona Walsh
Objectives of lecture
• Antibiotic discovery
• Time-line of currently prescribed antibiotics
• General principles of antimicrobial agents
• How antibiotics inhibit or kill bacteria
• Introduction to all antibiotic classes
Definitions
• Antibiotic is a naturally occurring
substance that inhibits or kills bacteria
• Antibacterial is a natural, semi-synthetic or
synthetic substance that inhibits bacteria
• Antimicrobial agent is a natural, semisynthetic or synthetic substance that
inhibits microbes
Antibiotic discovery
19th Century
Louis Pasteur
Robert Koch
Identified bacteria as causative agent of
disease. (Germ theory)
Now know what is causing disease, need to find out how to stop it.
1877 Pasteur
Soil bacteria injected into animals
made Anthrax harmless
1888 de Freudenreich
Isolated product from bacteria with
antibacterial properties. Toxic and
unstable.
Antibiotic discovery
20th Century
Erhlich
Worked with dyes and arsenicals worked
against Trypanosomes, very toxic.
1st antibacterial, only cured syphilis.
Domagk
Research on dyes.
1st synthetic antibacterial in clinical use.
Prontosil cured streptococcus diseases
in animals.
Active component: sulphonamide group
attached to dye.
Toxic.
Sulphonamide derivatives still used.
Less toxic.
Antibiotic discovery
20th Century
Fleming and
serendipity (1928)
Plates left on bench over weekend.
Staphylococcus colonies lysed/killed.
Fungi beside Staphylococcus.
Hypothesis: Fungi lysed Staph.
Unable to purify in large quantities.
No animal or human tests performed.
Antibiotic discovery
20th Century
Florey, Chain
Purified the penicillin from the fungus.
and Heatley (1939)
1940s (World War II) European and US cooperation led to
increased scale production of
penicillin.
Antibiotic discovery
20th century
Waksman (1943)
Isolated streptomycin from soil
bacteria Streptomyces.
Effective against Mycobacterium
tuberculosis and gram negatives.
Toxic antibiotic. Used until 1950s
when isoniazid used due to shorter
course of therapy.
1928
Discovery
Penicillin
1932
Sulphonamides
1939
Gramicidin
1930
1940
1942
1943
1945
1947
1949
1948
1952
Streptomycin & Bacitracin
Cephalosporins
Chloramphenicol & Chlorotetracycline
Neomycin
Trimethoprim
Oxytetracycline
Erythromycin
1956
1957
Vancomycin
Kanamycin
1961
1963
1964
1966
1967
1968
Nalidixic acid
Gentamicin
1971
1972
Tobramycin
Cephamycins & Minocycline
Fluoroquinolones
Oxazolidinones
1950
1960
Clindamycin
1970
1980s
1990s
General Principles
• Selective toxicity
– The essential property of an antimicrobial drug that equips it for
systemic use in treating infections is selective toxicity
– Drug must inhibit microorganism at lower concentrations than
those that produce toxic effects in humans
– No antibiotic is completely safe
General Principles
• Oral and Parental
– Oral antibiotics must be able to survive stomach acid
– Advantage: Ease and reduced cost
– Disadvantage: Circuitous route, antibiotic passes to lower bowel
– Parental antibiotics given by i.v.
– Advantage: Direct route to site of infection
– Disadvantage: Increased cost and need for qualified staff
General Principles
• Half-Lives
– The length of time it takes for the activity of the drug to reduce by
half
– Short half lives require frequent dosing
– Old antibiotics have short half lives
– New antibiotics may have half lives up to 33 hours
General Principles
• Broad and Narrow spectrum antimicrobials
– Broad spectrum antibiotics inhibit a wide range of bacteria
– Narrow spectrum antibiotics inhibit a narrow range of bacteria
– Broad spectrum desirable if infecting organism not yet identified
– Narrow spectrum preferable when organism has been identified
General Principles
• Bactericidal or bacteriostatic action
– Bactericidal antibiotics kill bacteria
– Bacteriostatic antibiotics inhibit the bacterial growth
– Bacteriostatic antibiotics may work as well as bactericidal
antibiotics if they sufficiently arrest the bacterial growth to enable
the immune system to eliminate the bacteria
General Principles
• Combinations of antibiotics
– Some antibiotics work better together than alone
– Combining 2 or more drugs may be required to prevent the
emergence of resistance e.g. tuberculosis
– Combinations should not be given when 1 drug would suffice
• Antagonistic effects
• No ability to adjust 1 drug concentration
Modes of action
Antimicrobial agents inhibit 5 essential
bacterial processes:
1.
2.
3.
4.
5.
Protein synthesis
Folic acid synthesis
DNA synthesis
RNA synthesis
Cell wall synthesis
1. Protein synthesis inhibitors
Protein synthesis
DNA
mRNA
transcription
Protein
translation
Ribosome is a protein factory in bacteria takes mRNA in and
produces proteins from them.
Bacterial ribosome has 2 parts:
–
–
30S binds to mRNA to translate mRNA into amino acids, which form
proteins
50S required for peptide elongation
3 phases from mRNA to protein
– Initiation
– Elongation
– Termination
Protein synthesis inhibitors
–
Aminoglycosides
–
Macrolides/Ketolides
–
Tetracyclines
–
Lincomycins
–
Chloramphenicol
–
Oxazolidinones
Protein synthesis inhibitors
• Bind irreversibly to ribosome
• Ribosome cannot bind to mRNA to form
amino acid chains (30S) or elongate the
chains to form proteins (50S)
• Disruptive effect on many essential
bacterial functions leading to cell death
2. Folic acid synthesis inhibitors
pterdine + para-amino benzoic acid
Dihydropteroate
synthetase
dihydropterate
Sulphamethoxazole
(Sulphonamides)
Structural analogues
of PABA
dihydrofolate
Dihydrofolate
reductase
tetrahydrofolate
Trimethoprim
(Diaminopyrimidines)
Binding
DNA/RNA
Reasons for combining
Trimethoprim and Sulphonamides
• There is synergy between the two drugs - the
combined effect is greater that the expected sum
of their activities
• Individually the drugs are bacteriostatic;
however, in combination they are bactericidal
• The use of two drugs will delay the emergence
of resistance
3. DNA synthesis inhibitors
• Enzymes required for DNA replication
• Topoisomerase II (DNA gyrase): GyrA and
GyrB
• Topoisomerase IV: ParC and ParE
• Quinolones interact/bind to the
topoisomerases, which stops DNA
replication e.g. nalidixic acid, ciprofloxacin
Action of fluoroquinolones
GyrA/
GyrB
DNA
ParC/
ParE
DNA gyrase
Topoisomerase IV
Quinolones
Cell death
DNA synthesis inhibitors
• Metronidazole
– Nitro group is reduced by bacterial enzyme
– Produces short-lived, highly cytotoxic free
radicals that disrupt the DNA
– Similar effect to UV radiation on cell DNA
4. RNA synthesis inhibitors
• Rifampicin
• Forms a stable complex with bacterial
DNA-dependent RNA polymerase
• Prevents chain initiation process of DNA
transcription
• Mammalian RNA synthesis not affected as
RNA polymerase is much less sensitive to
rifampicin
5. Cell wall synthesis inhibitors
– Vancomycin
– Bacitracin
– β-lactams
•
•
•
•
Penicillins
Cephalosporins
Carbapenems
Monobactams
– β-lactamase inhibitors
• Clavulanic acid
• Sulbactam
• Tazobactam
Action of Cell wall synthesis inhibitors
N-acetyl-glucosamine (NAG)
Phospho-enol pyruvate
Peptidoglycan formation
1. Building Blocks
N-acetyl-muramic acid (NAMA)
L-alanine
D-glutamic acid
L-lysine
NAMA
L-ala-D-glu-L-lys
D-ala-D-ala
NAMA
L-ala-D-glu-L-lys-D-ala-D-ala
D-ala
L-ala
Action of Cell wall synthesis inhibitors
NAMA
Lipid
carrier
Bacitracin
inhibits
L-ala-D-glu-L-lys-D-ala-D-ala
NAG
NAMA - NAG
L-ala-D-glu-L-lys-D-ala-D-ala
5 gly
Phospholipid
NAMA - NAG
L-ala-D-glu-L-lys-D-ala-D-ala
5 gly
Vancomycin
&Teicoplanin
binds, prevents
enzyme
polymerisation
Action of Cell wall synthesis inhibitors
Polymerisation
NAMA
NAG
NAMA
NAG NAMA
NAG NAMA
NAG NAMA
L-ala
L-ala
L-ala
L-ala
L-ala
D-glu
D-glu
D-glu
D-glu
D-glu
L-lys
5 gly
L-lys
5 gly
L-lys
5 gly
L-lys
5 gly
L-lys
D-ala
D-ala
D-ala
D-ala
D-ala
D-ala
D-ala
D-ala
D-ala
D-ala
NAG
5 gly
Action of Cell wall synthesis inhibitors
Transpeptidation
NAMA
NAMA
NAG
NAG
NAMA
L-ala
L-ala
L-ala
D-glu
D-glu
D-glu
D-ala
L-lys
D-ala
D-ala
D-ala
D-ala
L-lys
D-ala
L-lys
5 gly
NAG
5 gly
NAG
D-ala
L-lys
D-ala
D-ala
D-ala
L-lys
D-ala
L-lys
5 gly
D-ala
D-glu
D-glu
D-glu
L-ala
L-ala
L-ala
NAMA
NAG
NAMA
NAG
NAMA
b-lactams
resemble Dala-D-ala,
bind to
enzyme,
inhibit crosslinking
Penicillin Binding Proteins
Enzymes involved in cell wall formation
• Reseal cell as new peptidoglycan layers
added
• Penicillins bind to PBPs block enzyme
cross-linking chains
• Weak cell wall
• Build up osmotic pressure
• Lysis
Keynote points
• Recent history of antibiotic discovery
• General principles of antibiotic action
• 5 modes of action
• Examples of each