Antibiotics Lecture Notes PDF

BIO 418 ANTIBIOTICS Lecture I Seyhun Yurdugül Introduction  Content Outline Definition Historical background Classification of antibiotics Examples  Abbreviations m/o: microorganisms AB:Antibiotics DNA:Deoxyribonucleic acid APUA:Al...

BIO 418 ANTIBIOTICS Lecture I Seyhun Yurdugül Introduction  Content Outline Definition Historical background Classification of antibiotics Examples  Abbreviations m/o: microorganisms AB:Antibiotics DNA:Deoxyribonucleic acid APUA:Alliance for the Prudent Use of Antibiotics.  Definition in biological borders Any kind of agent bearing a chemical(synthetic) or biological origin; which is designed to eliminate microorganisms(m/o).  What are Antibiotics(AB)? Chemical compounds that are usually derived from substances produced by a mold or bacterium.  What are Antibiotics? Capable of killing or inhibit the growth of bacteria. Many antibiotics in use nowadays: synthetically made; or naturally produced (cited from APUA 1999)  What are Antibiotics? However, please note that antibiotics are only effective in: the treatment of bacterial infections, have absolutely no effect on viral infections. Historical Background ☻Since 1940, ☻ they are being considered as the most efficient; ☻ and desirable means of fighting various syndromes. ☻ Once being described as “magic bullets”; ☻ because they can eliminate bacteria; ☻ without bringing much side effects to the cells of treated individuals. ☻ they are overused, ☻ and the consequences: not “harmless”; ☻ but they are quite costly indeed.  Different situations of use Usage: not limited to fight against illnesses in human body. widely used throughout the ecosystem, such as veterinary practices (i.e. applied across to various sorts of animals), food-animal production, and even in selected plant species. In animals, they are used to prevent and cure infection.  Different situations of use Long-term exposure to low doses: the perfect formula for selection, increasing numbers of resistant bacteria in the treated animals; which may pass bacteria to caretakers and broadly, to people consuming undercooked meal.  Classification of antibiotics Several classification schemes for antibiotics, based on: bacterial spectrum (broad versus(vs.) narrow) route of administration (injectable vs. oral vs. topical) type of activity (bactericidal vs. bacteriostatic) chemical structure (the most useful classification)  Antibacterial agents Disinfectants Antiseptics Preservation agents Antibiotics  Disinfectants Usually too toxic, irritant or corrosive agents; to be applied to body surfaces or tissues, but suitable for removal of m/o’s from the equipment or the inanimate environment.  Antiseptics Include bacterial inhibitors; that are sufficiently free from the toxic effects, can be applied to body surfaces or exposed tissues; and are agents which should assist; and not impair the natural defence systems of the body.  Preservation agents Frequently added to pharmaceuticals; Cosmetics, and food products: to inhibit microbial contamination and proliferation  Different examples from antibiotics PENICILLINS : The oldest class of antibiotics. Have a common chemical structure which they share with the cephalosporins. The two groups are classed as: the beta-lactam antibiotics, and are generally bacteriocidal -that is: they kill bacteria rather than inhibiting growth.  Different examples from antibiotics CEPHALOSPORINS The usually preferred agents for surgical prophylaxis. Cefotaxime, ceftizoxime, ceftriaxone and others, cross the blood-brain barrier; and may be used to treat meningitis and encephalitis. Different examples from antibiotics FLUROQUINOLONES Synthetic antibacterial agents, not derived from bacteria. Can be readily interchanged with traditional antibiotics.  Bacteriocidal vs Bacteriostatic Bacteriocidal -that is, they kill the bacteria rather than inhibiting growth. Vice versa case: Bacteriostatic: they inhibit the growth of m/o rather than killing  Different examples-continued TETRACYCLINES They share a chemical structure that has four rings. They may be effective against a wide variety of microorganisms, including rickettsia and amoebic parasites.  Different examples-continued MACROLIDES Erythromycin, the prototype of this class, has a spectrum and use similar to penicillin. Newer members of the group, azithromycin and clarithromycin, are particularly useful for their high level of lung penetration. Clarithromycin: has been widely used to treat Helicobacter pylori infections, the cause of stomach ulcers.  Different examples-continued OTHERS include the aminoglycosides: particularly useful for their effectiveness in treating Pseudomonas aeruginosa infections; The Lincosamides, clindamycin and lincomycin, highly active against anaerobic pathogens.  Establishment steps of an infection by a pathogenic bacterium in man and animals Attachment to the epithelial surfaces of the respiratory, alimentary or urogenital tracts(1) Penetration of the epithelial surfaces by the pathogen(2) Interference with or evasion of the host defence mechanism(3) and multiplication(4) Damage of the host’s tissue(s) (5)  Main target of AB Prevention of the step (4) Either by killing the pathogens or slowing the growth to the point, where host defence mechanisms can clear the infection Attemps are trying to develop AB capable of bacterial attachment(step 1)  STRUCTURE OF BACTERIA Gram (+) Gram (-) Retaining the stain of iodine-crystal violet, when treated with organic solvents (e.g. alcohol or acetone):G(+) Staining response depends: primarily on the morphology, and composition of the bacterial cell wall.  Cell wall Surrounds the inner cytoplasmic membrane Maintains the shape of the cell; and protects the mechanically fragile cytoplasmic membrane from rupture G(-):inner region is peptidoglycan and lipopolysaccharide G(+): peptidoglycan and teichoic acid.  Capsules Discrete, tightly bound polysaccharide layers, distinct from the extracellular mucoid substance (glycocalyx or muco- exopolysaccharide) e.g. Pseudomonas aeruginosa. Glycocalyx structure provides a biofilm structure.  Peptidoglycan Mechanical stability of cell walls Polymer consisting of a disaccharide repeating unit of two different N-acetylated amino sugars, to one of which is attached a short peptide chain.  Peptidoglycan Sugars: N-acetyl glucosamine and N-acetyl muramic acid. Selective target for antibiotic action e.g. β- lactam.  Cytoplasmic membrane(CM) Lies directly outside the cytoplasm Acts as a selective permeability barrier, between the cytoplasm and the cell environment. Contains phospholipids and proteins, Provides the matrix: by which metabolism is linked to solute transport, flagellar movement and the generation of ATP.  Cytoplasmic Membrane Several antibiotics disrupt the CM by: -inducing leakage of intracellular constituents -inducing lysis -dissipating the proton motive force(pmf). Another target: Deoxyribonucleic acid(DNA) Bacterial chromosome: single, circular DNA No surrounding membrane and definite shape. Another target: Deoxyribonucleic acid(DNA) 18 different gene products participate directly in the replication of the E.coli chromosome. e.g.DNA gyrase, the target of quinolone antibiotics. Another e.g. Acridines  Spore structure Under extreme conditions. Spore formers: Bacillus, Clostridium, Sporolactobacillus, Sporosarcina,Desulfomaculum spp. ☻Germ cell (protoplast or core) and germ cell wall is surrounded by the cortex ☻Cortex is surrounded by spore coats  Spore structure-continued Bacterial spores: ☻ most resistant forms of all microbial forms to inactivation ☻ by chemical or physical agents (heat, radiation etc.) ☻ If normal conditions exist: Germination. G (+) G(-) (6A) Untreated and (6B) treated Bacillus subtilis spores. (Photos: courtesy of Dr. Alex Wekhof)  LITERATURE CITED Brock, T.D. Biology of Microorganisms, Eighth Edition, Prentice-Hall International Inc. 1998. Russell, A.D. and Chopra, I.; Understanding Antibacterial Action and Resistance, Ellis Horwood Ltd., Chichester, England, 1990.  BIO 418 Antibiotics Seyhun Yurdugül Lecture II Mode of action of antibiotics and their uptake into the bacteria  Outline Classification of antibiotics Different examples  Classification Nucleic acid synthesis inhibitors Protein synthesis inhibitors Peptidoglycan synthesis inhibitors Disruption of membrane integrity Inhibitors of Nucleic Acid Synthesis Growth mostly depends on DNA and RNA synthesis: Three main categories a- Interruption of nucleotide metabolism, by interference with nucleotide synthesis or nucleotide interconversion b-Interaction with DNA to form a complex; or causing strand breakage or removal of bases c-Antibiotics which directly inhibit enzymic processes in nucleic acid synthesis. Adenine(A) Thymine(T) Guanine(G) Cytosine(C)  Compounds(Antibiotics) that interrupt nucleotide metabolism Mainly interfere with the biosynthesis of tetrahydrofolic acid (THFA), a donor of one-carbon units at several stages: in purine and pyrimidine synthesis. e.g. Sulphonamides (Sulphamethoxazole) and 2, 4- diaminopyrimidine drugs (e. g. Trimethoprim)  Sulphonamides Sulphometoxazole and other sulphonamides: structural analogues of p-(4)-aminobenzoic acid (PABA), that bind more tightly to dihydropteorate synthase (DHPS) enzyme than PABA itself. Diaminopyrimidines (e. g. trimethoprim) completely inhibit bacterial dihydrofolate reductase(DHFR).  Antibiotics interfering enzymes in nucleic acid synthesis Inhibitors of RNA polymerase e. g. Rifampicin group, a semi synthetic group, binds and specifically inhibits bacterial DNA dependent RNA polymerase, by inhibiting the initiation process Antibiotics interfering enzymes in nucleic acid synthesis ♣ Rifampicin group shows: ♣ no effect on nuclear or mitochondrial DNA dependent RNA polymerase(Selective action)  Antibiotics interfering enzymes in nucleic acid synthesis Inhibitors of DNA gyrase The enzyme involved in bacterial DNA replication. In other words called DNA topoisomerase II Quinolone group antibiotics have particular interest in inhibition of DNA gyrase. Overlap of the novobiocin (white) and the ADPNP (blue) binding sites. Overlap of the cyclothialidine (yellow) and the ADPNP (blue) binding sites.  Escherichia coli(E.coli) DNA gyrase Four subunits Two gyrase A subunits (each MW: 97000) Two gyrase B subunits (each MW: 90000)  Quinolones Nalidixic acid, an 8-aza-4-quinolone (inhibitors of DNA gyrase) Other e.g from group: Norfloxacin, enoxacin, ofloxacin and ciprofloxacin.  Quinolones Inhibition against a wide spectrum of m/o Bind predominantly to A subunit of DNA gyrase, impairs both A & B subunit function, induces SOS response.  LITERATURE CITED Brock, T.D. Biology of Microorganisms, Eighth Edition, Prentice-Hall International Inc. 1998. Russell, A.D. and Chopra, I.; Understanding Antibacterial Action and Resistance, Ellis Horwood Ltd., Chichester, England, 1990.  BIO 418 Antibiotics by Seyhun Yurdugül Lecture III Inhibitors of Protein Synthesis  Outline Basic information about protein synthesis Selectivity mechanism for antibiotics Various examples  Abbreviations S:Svedberg unit mRNA:messenger RNA tRNA:transfer RNA DNA:Deoxyribonucleic acid. Information about protein synthesis: next figures  Summary of Protein Synthesis The ribosome binds to the mRNA; at the start codon (AUG); that is recognized only by the initiator tRNA.  Summary of Protein Synthesis The ribosome proceeds to the elongation phase of protein synthesis. During this stage, complexes, composed of an amino acid linked to tRNA, sequentially bind to the appropriate codon in mRNA; by forming complementary base pairs with the tRNA anticodon.  Summary of Protein Synthesis-contd The ribosome moves from codon to codon along the mRNA. Amino acids are added one by one, translated into polypeptidic sequences dictated by DNA, and represented by mRNA.  Summary of Protein Synthesis-contd At the end, a release factor binds to the stop codon, terminating translation, and releasing the complete polypeptide from the ribosome. One specific amino acid can correspond to more than one codon. The genetic code: said to be degenerate.  Selectivity of these AB Specifically bind to bacterial ribosomes rather than mammalian ribosomes e.g. E.coli ribosome, 70 S composed of 30 S & 50 S subunits Carry important functional sites, like the aminoacyl tRNA, and peptidyl tRNA binding sites  Aminoglycosides includes at least eight drugs: amikacin, gentamicin, kanamycin, neomycin, netilmicin, paromomycin, streptomycin, and tobramycin, having the same basic chemical structure.  Aminoglycosides primarily used to combat infections due to aerobic, Gram-negative bacteria. Bacteria that can successfully be combated with : Pseudomonas, Acinetobacter, and Enterobacter species, also effective against Mycobacteria.  Streptomycin the first aminoglycoside, isolated from Streptomyces griseus in the mid-1940s. very effective against tuberculosis.  Streptomycin one of the main drawbacks to streptomycin : is its toxicity, especially to cells in the inner and middle ear and the kidney. some strains of tuberculosis: are resistant to treatment with streptomycin. Action mechanism of Streptomycin bactericidal ability has not been fully explained. the drug attaches to a bacterial cell wall; and is drawn into the cell via channels made up of the protein, porin. inside the cell, the aminoglycoside attaches to the cell's ribosomes.  Action mechanism of Streptomycin This attachment either a) shuts down protein production or b) causes the cell to produce abnormal, ineffective proteins. so bacterial cell cannot survive.  Other aminoglycosides Kanamycin Bind to the 50S subunit of the ribosome and prevent translation; from beginning by distorting the ribosome so it can no longer carry out its main functions.  Other aminoglycosides Although most of the AB from this group are lethal due to the inability, to make essential proteins, kanamycin can be lethal: when the whole of 30S tRNA subunits begin to be disrupted. Actually a combination of three antibiotics that work together.  Tetracyclines also target ribosomes. They differ from the aminoglycosides in that: they bind to a different site on the ribosome (70S and 80S).  Chloramphenicol inhibits a class of enzymes: that aid in growing the polypeptide chain during protein synthesis. binds to the 70S subunit of the ribosome.  Macrolide antibiotics prevent elongation of the growing protein, translocation of the ribosome along the mRNA, or both. E.g. Erythromycin Binds to a specific site on the ribosome (50S).  LITERATURE CITED: Brock, T.D. Biology of Microorganisms, Eighth Edition, Prentice-Hall International Inc. 1998. Russell, A.D. and Chopra, I.; Understanding Antibacterial Action and Resistance, Ellis Horwood Ltd., Chichester, England, 1990.  BIO 418 Antibiotics by Seyhun Yurdugül Lecture 4 Peptidoglycan Synthesis Inhibitors  Content Outline Peptidoglycan Structure Peptidoglycan Synthesis Inhibitors Beta-lactams  CONTENT OUTLINE General Information About Vancomycin History and Background Mechanism of Action Mechanism of Resistance Pharmacokinetics Clinical Uses Side Effects Alternatives of Vancomycin for Less Serious MRSA Infections  Abbreviations GIT:gastrointestinal tract IV:Intravenous kD:kiloDalton mcg:microgram mg:miligram  Peptidoglycan Synthesis Inhibitors ♣ Most bacteria have a cell wall, containing a special polymer called peptidoglycan. ♣ Over the cell membrane: a shift of peptidoglycan; and other polymers including teichoic and teichuronic acids. ♣ Peptidoglycan gives a certain rigidity to the cell wall; and gives the cell mechanical strength.  Peptidoglycan Synthesis Inhibitors ♣The bacterial cell wall is a unique biopolymer, it contains both D- and L-amino acids. ♣ Its basic structure is a carbohydrate backbone of alternating units of N-acetyl glucosamine and N-acetyl muramic acid.  N-acetyl muramic acid ♣ The N-acetyl muramic acid residues are cross- linked with oligopeptides. ♣ The terminal peptide: D-alanine although other amino acids are present as D- isomers. This is the only known biological molecule: contains D-amino acids, and it is the target of numerous antibacterial antibiotics; e.g. penicillin. Peptidoglycan of Staphylococcus aureus Peptidoglycan Synthesis of Bacteria Takes place in three major stages Stage 1-Synthesis of precursors in the cytoplasm St.2 -Transfer of precursors to a lipid carrier molecule(undecaprenyl-phosphate), which transports them across the cytoplasmic molecule St. 3-Insertion of glycan units into the cell wall attachment, by transpeptidation and further final maturation steps  Stage 1 inhibitors D-cycloserine Blocks peptidoglycan synthesis by competitive inhibition of two enzymes: alanine racemase and D-alanyl D-alanine synthetase.  Stage 2 inhibitors Bacitracin: complexes with the membrane bound pyrophosphate form of the undecaprenyl(C55- isoprenyl)lipid carrier molecule, that remains after the disaccharide pentapeptide unit has been transferred to peptidoglycan chain.  Stage 3 inhibitors Glycopeptide antibiotics Vancomycin, ristocetin and teicoplanin E.g vancomycin combines with peptidoglycan substrate; rather than an enzyme. Peptidoglycan of Escherichia coli  The Most Important Inhibitor of Stage 3 Penicillin (β-lactam) inhibitors  Beta-lactam antibiotics Penicillin nucleus Cephalosporin nucleus  Action of beta-lactam antibiotics ♣ Inhibition of final stages of assembly of the peptidoglycan ♣ must penetrate the cell wall and operate in the periplasmic space ♣ bind to 'penicillin-binding proteins' in the outer leaf of the cell membrane ♣ these are the enzymes responsible for the final stages of assembly of the peptidoglycan molecule  Structure of β-Lactam Rings The 3-D structure of a beta-lactam closely resembles: the D-alanyl-D-alanine end of the peptide in peptidoglycan; just before final assembly: penicillin binds into the active sites of several of the enzymes; which normally work on D-alanyl-D-alanine; and blocks their action.  Structure of β-Lactam Rings this prevents synthesis of the final stable peptidoglycan molecule this molecule is essential to the stability of the bacterial cell wall, and so the affected cell disintegrates In Benzyl penicillin molecule, two D-alanine molecules: in the same orientation. Benzyl penicillin molecule An example: Vancomycin  GENERAL INFORMATION ABOUT VANCOMYCIN Vancomycin Class: Glycopeptide antibiotic Type:Stage III inhibitor -Synthetic antibiotic Formula:1.0 g vancomycin base, Flacon 1.0g or 550 mg vancomycin base, 500mg Flacon Radical Group:Hydrochloric Acid Indications: -Effective against methicillin-resistant Staphylococcus(Beta-Lactam resistant) -Penicillin allergy -Cephalosporins and other antibiotics resistant microorganisms -HCl : Staphylococcal endocarditin -Use with rifampin or aminoglycosides or use with both of them. HISTORY AND BACKGROUND Vancomycin 1.5kD glycopeptide. Introduced in hospitals ~40 years ago when strains of bacteria exhibited penicillin resistance. Vancomycin resistant enterococci (VRE) appeared in 1987.  Mechanism of action (MOA) Inhibits bacterial cell wall synthesis. -Vancomycin binds to the D-alanyl-D-alanine residues of the peptidoglycan monomers. Cross links are not formed: so the peptidoglycan chains only form a weak cell wall. Intense osmotic pressure ruptures the cell.  Spectrum of action: Gram positive organisms Including: Listeria, Rhodococcus, Peptostreptococcus Bacteriostatic against Enterococcus  MECHANISM OF RESISTANCE Mechanism of resistance: – Enterococcus: Van A – E Peptidoglycan precursor has decreased affinity for vancomycin – D-ala-D-ala replaced by D-ala-D-lac. – Staphylococcus aureus: Vancomycin induced Staphylococcus aureus(VISA) isolates: – Increased amount of precursor with decreased affinity – Thicker cell wall hVISA: heterogenous vancomycin induced Staphylococcus aureus(VISA) bacterial population  binding to the terminal D-alanyl-D-alanine residues → prevents crosslinking of the peptidoglycan component in the cell wall of G(+) organisms Inhibits bacterial growth, eventually leading to death. Enterococcus faecalis  VRE D-alanyl-D-alanine residue ↓ D-alanyl-D-lactate moiety vancomycin cannot bind to this peptide NOT effective in G(-) bacteria  Mechanism Vancomycin binds with high affinity to the terminal D-Ala-D-Ala through five hydrogen bonds.  PHARMACOKINETICS Not absorbed from GIT( given oraly in antibiotic- associated colitis, esp. if caused by C. difficile ) It is given intravenously(IV) Widely distributed ( including CSF in inflammation ) Only 10 % bound to plasma protein Eliminated entirely by glomerular filtration Accumulate in renal impairement Half- life approximately 8 hours  CLINICAL USES Serious Staphylococcus infections caused by methicillin resistant Staphylococcus aureus(MRSA) or MRSE(methicillin resistant Staphylococcus epidermitis). Prophylaxis for major surgical procedures in hospitals with high rates of infections due to MRSA or MRSE Antibiotics-associated colitis – metronidazole is preferred ) Vancomycin- resistant Staphylococcus use Daptomycin  CLINICAL USES Dose: – Based on total body weight and renal function – 15 – 20 mg/kg – Normal renal function: q 12 dosing Goal through concentrations: – 10 – 15 mcg/mL: bacteremia, skin and soft tissue infections – 15 – 20 mcg/mL: osteomyelitis, meningitis, pneumonia  SIDE EFFECTS Fever, rashes Thrombophlebitis at the site of injection Ototoxicity & nephrotoxicity ( High concentration ) Hypotension and red man ( red neck ) syndrome( rapid infusion ) ALTERNATIVES TO VANCOMYCIN FOR LESS SERIOUS MRSA INFECTIONS Skin and Soft Tissue – Trimethoprim-sulfa (TMP/SMX) – Doxycyline/minocycline – Clindamycin if no inducible resistance – NOT LEVOFLOXACIN! ALTERNATIVES TO VANCOMYCIN FOR LESS SERIOUS MRSA INFECTIONS Urinary Tract Infections (always rule out bacteremia before Rx) TMP/SMX – Not doxycycline or minocycline or linezolid Not adequate urinary concentration Plain tetracycline if susceptible (94%) – Perhaps, fluoroquinolones if susceptible in vitro  Daptomycin Linezolid Tigecycline (not for bacteremia) Ceftobiprole (not yet FDA approved) VRE  LITERATURE CITED 1. Brock, T.D. Biology of Microorganisms, Eighth Edition, Prentice-Hall International Inc. 1998. 2. Russell, A.D. and Chopra, I.; Understanding Antibacterial Action and Resistance, Ellis Horwood Ltd., Chichester, England, 1990. 3. Rybak MJ et al. Vancomycin Therapeutic Guidelines: A Summary of Consensus Recommendations. Clin. Inf. Dis., 2009;49:325-327. 4. Rybak MJ et al. Therapeutic monitoring of vancomycin in adult patients: A consensus review. Am J Health-Syst. Pharm., 2009;66:82-98 5. Rybak MJ The Pharmacokinetic and Pharmacodynamic Properties of Vancomycin. Clin. Inf. Dis., 2006:42:S35-39 6. Stryjewski ME et al. Use of Vancomycin or First-Generation Cephalosporins for the Treatment of Hemodialysis-Dependent Patients with Methicillin-Susceptible Staphylococcus aureus Bacteremia. Clin. Inf. Dis., 2007; 44:190-196. 7. www.ilacrehberi.com (cited 30.07.2011) 8. MedicineNet.com (cited 30.07.2011) 9. www.rxlist.com (cited 30.07.2011)  BIO 418 Antibiotics by Seyhun Yurdugül Lecture 5 Penicillin-Binding Proteins (PBPs)  Content Outline Types of Penicillin Binding Proteins Targets of Beta-lactams Lytic and non-lytic death Transport mechanism for uptake  Abbreviations PBP: penicillin binding protein. E.coli: Escherichia coli  Introduction Identification of more than one penicillin sensitive reaction; in the cell free peptidoglycan synthetic systems: implied that several proteins(enzymes) are capable of : interacting with penicillin(and other beta lactam antibiotics) might be present in any bacterial species -these are penicillin-binding proteins (PBPs)  Types and some properties of PBPs Several (usually at least four) are present in bacterial species PBPs are designated numerically e.g. one- seven in E.coli K12 are minor components of the cell membrane  Types and some properties of PBPs can catalyze: transpeptidase, carboxypeptidase, and endopeptidase reactions (as they are enzymes in nature)  E.coli PBPs The most extensively studied set of PBPs. Essentially divided into two groups a)High molecular weight E.coli PBPs b)Low molecular weight E.coli PBPs  High molecular weight E.coli PBPs PBPs (1-3) responsible for net synthesis of peptidoglycan in vivo  High molecular weight E.coli PBPs catalyze both the polymerization of the disaccharide units into glycan chains(transglycosylation), and also the cross linking of their pentapeptide side chains(transpeptidation)  Special conditions Inhibition of the transpeptidase activity of the high molecular weight PBPs of E.coli by β-lactam antibiotics, eventually leads to cell death.  Special conditions-II Inhibition of PBPs 1A & 1B leads to rapid cell lysis, Inhibition of PBP 2 leads to growth as osmotically stable spherical; or ovoid forms, and Inhibition of PBP 3 to filamentation.  Low molecular weight E.coli PBPs PBPs 4-6 PBP 4 has the activity of D-alanine carboxypeptidase and endopeptidase PBPs 5 and 6 catalyze also D-alanine carboxypeptidase reaction. PBP 7: Enzymatic activity and function is not identified.  PBPs are targets for β-lactams binding of a β-lactam: leads to inactivation of transpeptidase activity: potentially lethal to bacteria. Rapid lysis of bacteria results from inactivation of PBPs 1A and 1B  Other main targets Peptidoglycan carboxypeptidases and transpeptidases: the main target of β-lactams Their normal substrate: D-alanyl D-alanine moiety of the pentapeptide, but penicillin and other β-lactams: will bind covalently to the same group in the PBPs that normal substrate binds.  Other mechanisms for inhibition of peptidoglycan synthesis Lytic death Non-lytic death  Lytic death Peptidoglycan degradative enzymes (autolysins) of some bacteria Enzymes that hydrolyse bonds in the glycan or peptide side chains of peptidoglycan Together with any kind of antibiotics like penicillin: facilitate cell lysis.  Non-lytic death Observed in bacteria having no autolytic activity Seen as ‘peptidoglycan nicking’, and a very limited amount of wall damage Uptake of antibiotics by the bacteria In order to reach the target sites the antibiotic molecules has to cross either one or two membranes Mainly three methods a) Self promoted uptake b) Passive diffusion c) Active transport  Self-promoted uptake Involves destabilization, and disorganization of the outer membrane, as a result of displacement of divalent cations Mostly carried out by polycationic antibiotics. The mechanism is not yet been fully understood.  Polycationic group Main representatives are A)Aminoglycosides B)Polymyxins  Passive diffusion Can be also called as simple diffusion, occurs primarily through the water field pores(so-called ‘porins’)  Passive diffusion It is mainly influenced by the molecular size, charge and lipophilic properties of the permeating molecule.  Active transport Involves specific carrier proteins that are coupled to an energy source. When metabolic energy is available, antibiotics can be accumulated within the cell against a concentration gradient. Energy arises from ATP, or the proton motive force(PMF)  Several examples for uptake E.g. 1 Quinolones These antibiotics mainly cross, for ex. E.coli outer membrane by specific molecules (Omp C and OmpF porins, cause nucleic acid inhibition.  Several examples for uptake E.g. 2 Aminoglycosides Cross both inner and outer membranes: by two phases, EIP(energy independent) and EDP(energy dependent) 1 and 2., as target: ribosome  Several examples for uptake β-lactam inhibitors As their target is peptidoglycan; they have to cross the outer membrane, by passive diffusion through porin channels.  Example to β-lactam inhibitors Polymyxins (membrane integrity inhibitors) Interacts with the cytoplasmic membrane, to cause gross disorganization of the structure  LITERATURE CITED Brock, T.D. Biology of Microorganisms, Eighth Edition, Prentice-Hall International Inc. 1998. Russell, A.D. and Chopra, I.; Understanding Antibacterial Action and Resistance, Ellis Horwood Ltd., Chichester, England, 1990.