Cellular processes in Protein Synthesis va sd A DRA ein” DNA ain @ a5 KK m Hf ie gall RNA AN". iin « gm Protein o=)= 0) Translation @ Protein GP. GTAL angusProtein synthesis and is Inhibition a) The bacterial ribosome isa 70S particle made up ofa 30S subunit anda 505 subunit j The 30S subunit binds messenger RNA (mRNA) and initiates protein synthesis, The 50S subunit combines with the 305 subunit-mRNA complex to forma ribosome, then . ; binds aminoacy transfer RNA (CRNA) and catalyzes the buldng ofthe protein chai, = S74 d ¢ U ¢ ¢ A nal bt A There are two main binding sites for the tRNA molecules. The peptiyl site Psite) cw c 4 A binds the tRNA bearing the peptide cain, The acceptor aminoacl ste (Ate) binds. jp)” the tRNA Dearing the next aminoacid, to which the peptide chain wl be transferred, nH | €} The rbsomes of ekayticcel are bigger 0), consiting of 6S and AS subunits, CuG = C) f)The agents described inthis secton al inhibit protein synthesis by binding to ribosomes, "f° and inhibiting diferent stages ofthe translation proces, a OG Ch ) Toxicity is due to either cifferent ciffusion rates through the cell barriers of bacterial rn th Versus mammalian cells or toa difference between the ribosomal target structures,Sal ribosomal subunit +5’ end of mRNA start codon {methionine}; there large ribosomal subunit , and {RNA carying amino acid join it \ Other RNAwithanticodon matching (complementary) ss bie to MRNA codon fal into place in the ribosome : a First RA drops its aminoacid, breaks and leaves to Dick up another amino acid the second moves over to make room for another tRNA ere {RiAafter tRNA dropoff amino acid and they fom a chain of aminoacid linked by peptide bond When ribosome reaches stop codon (Arginine), it releases finished polypentideStages at which Antibacterial inhibit translation Orazolicinones bind to 50 subunit Amino acid Chloramphenicol Macrolides Growing. | Jana blocks peptide block protein chain transfer translocation be | \ ’ ! | Ribosome AA ’ > > a4 ccs Niscelaneous «Streptomycin «Onytetracytine «ytiomen «Linezolid -Coanpenial «Neomycin Chlrtetracycine «Clarithromycin Sutealid Cinycn sRanamycin shinocytine «Azithromycin «Dovycyctine 9.6TAL. pangs 1 nisi Aminoghycosides blocks tRNA block translocation bindingAminoglycosides * Streptomycin was isolated from the soll microorganism actinomycetes Streptomyces griseus in 1944, + Aiinoglycsides basic in nature; pory absorbed in gastrointestinal tract. * Work best in slightly alkaline conditions, At pt 7.4, they have a positive charge that is beneficial activty by aiding absorption through the cuter membrane of Gram-negative bacteria, Also as sulfa salts highly soluble in water, * A ioic interaction takes place with various negatively charged groups on the outer surface ofthe cl membrane which displaces magnesium and calcium ions, These ions normally ac as bridges between lippalysaccheides, and their displacement results in rearrangement of cell membrane components t produce pores through which an aminoglycoside can pas, Nechanism of Action: * The binding is specfclly to the 305 ribosomal subunit and prevents the movement (translocation) ofthe ribosome along mRHAso tat the tile code on mRNA can no longer be red (misreacng), "Accumulation of released peptide from ribosomal side at cll membrane increases cel permeability resting in an even greater uptake ofthe drug,t * Wy ALY HH 4 sariowge ° nA ic ing | Stepiing ne i tH Kanamycin B oH 0 HO’ WA "als 0 oy $Aminosigar AVAy i AN ay, Pani ‘2dennysrepaning Tobranycin OH 0 HEA Ct my HO, . ‘ oe 0sugar rm a, aga 2deoxsteptaning Galant I tH 2 4eonsteptine sa na Ale Ribose HN, Ni ASC, ty re Lneosanine Pe : 7 eon teplaine a HN 0 ey NeomycinB AmiteinStructure Activity Relationship (SAR + 76 amino groups optimal for activity: Replacement wih Oh decreases activity eg, KanamycinAand C * Hetylation of C and NH, confers metabolic resistance; 6.9, gentamycin OH "4 t oH mA ” + }'or4' OH group nonessential eg, Tobramycin md HO ; uu WALI Ay 4-5 unsaturation (Netimicin: resist phosphorylated enzyme i 5 3 | *+ etait alan of ann gtup eg, aac retains activity wth enhanced stability against bacterial deactivating enzymes ¥ J-amino non-essential for activity, aia ata vit impacting activityO Deoxystreptamine-containing aminoslycosides ciffer quantitatively ftom strep a reading at lower concentration than those required to prevent initiation of protein o Jectinomycin prevents the initiation of protein synthesis but apparently does not cause y = noe coside-inactvatng bacterial enzymes include a) aminoacetytranfeases (HH, 3-NH,) ) phosphotranserases (3'-On,2”.Ot) (c)nucleotiayl transferases (40H) Significantly contributing to drug resistance, ¥ Aminolyosides are bactericidal athe than bacteristatic ands thous hath act ma 0 ther effects both onthe ribosomes andthe outer cell membrane, ~ Jecause the ribosomes in human cels are diferent in structure from those in bacterial cell, much lower binding affinity forthe aminoelycosides, which explains the selectivity of tee Y Usvaly administered as systematic route (Streptomycin), Kanamycin, neomycin, gentamycin ae use topical, na ’Clinical aplications of aminoglycosides a) Treatment of serous systemic infections caused by aerobic Gram-negative bacli ie, Urinary tract infection, diarrhea, peritonitis, meningitis, septicemia, biliary tract infection et. b) Aerobic Gram-negative and Gram-postive cocci (with the excention of staphylococc) tend to be less sensitive than 6-actams, ¢) Streptomycin is the most effective of the group forthe cHemotherapy of TB, €) Aminogycoside and B-actam antibiotics exert a synergistic action in vivo against some bacterial stains, For example, carbenicilin and gentamicin, Damage to the cel wall caused by the f-lactam antibiotic is believed to increase penetration of the aminoglycoside into the bacterial cell, Hence, effective against entercoccal endocarditis, Hence, categorized as broad spectrum. The two antibiotic types should not be combined inthe same solution because they are chemically incompatible, €} Not absorbed in GIT; Used to clean infection in GIT Adverse effect: « undesrable side effects, particularly ottovicity and nephrotoycty, have restricted systrmc use to serous infections o infections cased by bacterial strains resistant to other agentsTetracyclines a) The tetracyclines are obtained by fermentation procedures from Streptomyces spp, or by chemical transformation ofthe natural products, Oe b} Its derivative ofan octacydro naphthacene, a hydrocarbon sytem that comprises four annulatedsismembered rings, ie, Removd ¢) Stereocheristry: Carbon atoms 4, 4a, 5, 5a, 6, and {2a are potently chiral, 4 Q Chxytetracycline and doxycycline, each with a So-yroxyl substituent, have Ci sxasyrmetric centers th ote, lacking chai at, have only ie, °ehtapthacne k Q k # th Of Wh, oo Tetracycline Chlortetracyclne Onytetracyctine Doxycycline Ninocycined) The tetracyclines are amphoteric compounds, forming salt with elther acids or Solutions, these substances exist mainly a5 aviterions, The acid salts, which are protonation ofthe amino group on C-4, exist as crystalline compounds that are very sl ¢) Hydrochloride salts are used most commonly for ora administration and usvally are because they ae bitter. Basic salts are not stable in an aqueous solution f} The unusual structural groupings in the tetracyclines produce three 9 | Acidity constants (pKa) in aqueous solution ofthe acid als, \ Me | OH 0 of 0 Tetracycline hydrochloride WKa®T4 thee atianies Pha *83Degradation a) Tetracycines undergo epimerization at C4 in W solutions of intermediate pH range, These isomers AH Exineton MH mm are called epitetrayctnes, They exhibit much less $4 activity than the “natural” isomers, thus accounting , 00 ; Tetracycline for the decreased therapeutic value of aged —Agive solutions, = rong acids attack tetracyclines witha hydroxyl group on C-6, causing a loss in activity through S = = = = Ss S =. = cs = & a. Ss =sS.- S 3 a > = = Ss = s Ss s s a => a = = =. Pa s \/ HO) CH yt Stong adic LAA OH degradation VAN / ay NY: a ny! ‘ ( 010 Of 40 OHOHO 0 0 Tetracycline Anhydroetracycine Resonance and energetical favoured Less aclve¢) Bases promote a reaction between the O-hydroxyl group and the Ketone group causing the Dond between the {1 and {fa atoms to cleave, forming the lactone inactive isotetracyclne, €) These unfavorable reactions stimulated research that led to the development of the longer acting compounds b-ceoxytetracycin, methacyclne, doxycycline, and minocycln Stongbasic degradation Tetacycine Iotetracyclne Inactive* Stable chelate cor calcium, magnestu accounting fort Reactions of Tetracyclines plexes are formed by the tetracyclines with many metals, incluing , and ion, Such chelates are usually very insoluble in wate, impaired absorption of most if ot all tetracyclines inthe presence of milk; calcium, magnesium-, and aluminum:-containing antacics: and iron salts, * Soluble alkalinizrs, , Such as sodium bicarbonate, also decrease the Cl absorption of the tetracyclines, Deprotonation of tetracyclines to more ionic species and the observed instability ofthese products in alkane solutions may account fr ths observation, * The affinity of tetracyclines for calcium causes them to be incorporated into nevly forming Dones and these antibiotics i teeth as tetracycline-calcium orthophosphate complexes, Deposits of teeth cause a yellow discoloration that darkens (a phtocherical reaction) overtime, * Tetracyclines are distributed into the milk of lactating mothers and vill cros the placental barr int the fetus, CS? GTN campusNechanism of Action and Resistance Y Tetracyclines are spectic inhibitor of bacterial protetn synthesis, They bind to the 30S ribosomal and, thereby, prevent the binding of aminoacyl tRNA tothe mRiA-ribosome comple, > Tetracylins also bind to mammalian ribosomes but with lover affinities, and they apparently d achieve sufficient intracellular concentrations to interfere with protein synthesis, > Three biochemically cistinct mechanisms of resistance to tetracyclines have been described in bacteria * efflur mediated by transmembrane-spannig, actve-transport proteins * ribosomal protection, in which the bacterial protein synthess apparatus is rendered resistant "enzymatic oxidation, Clinical applications 0 The tetracyclines are broad-spectrum antibacterial agents, 0 They are active against spirochetes, mycoplasma, rcketsiae, and chlamydia, {2 Ther bacterosatic action, However, isa disadvantage in the treatment of life-threatening infection aS septicemia, endocardts, and meningts; the aminoslycosides and/or cephalosporins usvll prefered for Gram-negative andthe pencilns for Gram-positive infections, {1 Because of incomplete absorption and kiting natural bacterial flora of the intestine, tetracyctin induce supernfectons caused by the pathogenic yeast Candida albicans,Structure-Activity Relationships «Strongly electron withdrawing ctora group (chortetrayctne), ston electron donating group meth ano Winaycie) enhance actty ‘anplar group ctor, cimethy enhances GIT absorption despite loner water olubity «CL tycron, met group not necessary foractiy eg, -demethy-b-deotetracycing, metacycine active «C6 ceony substituents mare ipophilic eg, doxycycline, minocycline ond absorption in GI, higher protein binding, decreased renal clearance in contrast to more water-soluble compounds tetracycline and onjtetraccle, sAratization leds to inactivity eg, anhyotetrayctne «Stydro gu, a in ntetracytne and dnyytn, may influence pharmacokinetic properties (ality) «Araatzaton or inrductin of double bond reduces atty «Dinet-aino grou atthe C4 ust hav c orientation and is essential 4-ptetracytnes very much es active «The ended tricaron-metane sytem t (1 toC-3 mst be intact for good actty «Replacement ofthe aide a (2 with other functions (aldehyde or nitrile} reduces or abolishes acti Arvin lylaton of the amide trogen enhances solubility e.g, Roitetrayctne Tetracycing « Essential for activity along wth blue coloured SyOups (1,2, 3,4 10,1, ta, and 4, reresentng the hydrophilic Faces essential for actnty «Pasitin (5, 5a 6,7, 8, $ represent hyropobic face Changes inact lesson act, more on pharmacokinetics ra «CisA/B-ing fusion wth a hycroyl grup a Cais aparently bo essential, + Controls pharmacokinetics fain of 1 Hgts tityNacrolide antibiotics * Hacrotides are bacteriostatic agents isolated from the soll actinomycetes Streptomyces s + The structure consists ofa 14 to t6-membered macrocyclic lactone rng with a sugar anda + Because erythromycin and chloramphenicol bind tothe same resion ofthe ribosome, they are ' Entonyc is unstable to stomach acids; which led to te development ofa semisynthetic ana " y_ Anno sua Dessain) Aino sugar (Desosanin) Hos | \ Wy 64 NW 00 / "Hy 0 0 Laclone OH 0H Suga (ae) ws... Sugar (Cladinose) f Erythromycin X= H Aaithromycin Clarithromycin X= OCH, (SP.GTAL 1 canes* Erythromycin canbe taken orally in a tablet form. The formulation of the tablet invol is designed to protect the tablet during its passage through the stomach, but which i reaches the intestines (enterosoluble), Acid mediated degradation Product * The acid sensitivity of erythromycin is cue to the presence of a ketone and two alcohol gro are setup forthe aci-catlyzed intramolecular formation f a ketal Enythomycin Inactive metaboliteStructure-activity Relationship a) The sugar residues are important for activity 0) Protection of acid senstve hydroxy group by converting to ethoxy enhances stability improves oral absorption e.g, Clarithromycin, ¢) Replacement of acid sensitive carbonyl group (Azithromycin) further improves pharmaco €) Increment in ring size to 16 membered rng further prevents acid sensithty and improves a Tlthromycin that contains a keto and a carbamate group in place of replaceable sugar unit hydroxyl respectively N I+ r Protection of acid ble OH Acid sensve groups CH) N Wasting of acd tile OH roe > Reylceable Telithromycin ErythromycinClinical applications of Macrolide antibiotics 1 Treatment of Respiratory infections ¥ Mycoplasma pneumonia ¥ Rhinosinusts (inflammation ofthe nasal cavity and paranasl sinuses) ¥ Pertussis (highly contasious respiratory tract infection} ¥ Diphtheria (nflammation of the mucous membranes ofthe throat} ¥ Respiratory infections caused by Chlamycia trachomatis ¥ Bronchiectasis(a condition where the bronchial tubes of lungs are permanently damaged) ¥ Cystic fibrosis (a csorder that produces abnormally hick mucus leading to blockage of bch in ¥ Pharynts (inflammation of he back of he trot, often refered tos sore throat) ¥ Tonslitis(nflammatin of the tonsils) ¥ Acute exacerbation of COPD (chronic obstructive pulmonary csease) JChloramphenicol a) Chloramphenicol is aevorotatory broad-spectrum antibiotic produced from Streptomyc b) It has to asymmetric centers, of which only the RR isomer is active, ¢) Chloramphenicol binds to the 50S subunit of ribosomes and appear to act by inhibin ribosomes along mRNA, probably by inhibiting the pentidl transferase reaction (peptide ext €) Since it bind to the same region as macrolides and lincosaides, these crugs cannot be use Structure Activity Relationship The nitro group and both alcool groups are involved in binding interactions, + The cchloroacetamide group is also important, but can be replaced by othe electronegative groups, + Aromatic ring could be different substituted 9 ‘ without deterioration of activity, Chidation of OF produces inactive derivative, e | chloramphenicol 30H canbe eseried for better pharmacokinetics,Degradation Hydrolysis of chloramphenicol with acid produces dichloroacetic acid together with active base while in alkali hydrolysis generates , o- cydrxy mgt f Achy oe we i Tita O bap Vv 0 we “A ok | a. chornphnicol ase udahgs 40 " We" @, i dry vi * Chloramphenicol is quite toc andthe nitro substituent i thought to be responsible for ts, + Hain metabolic pathway involves formation of the 3-0-slucuraide, reduction ofthe p-nitro aromatic amine, hydrolysis ofthe amide, and hydrolysis of the a-chloracetamido group, a * ‘Strains of certain bacterial species are resistant to chloramphenicol by virtue of the a chloramphenicol acetytransferase, an enzyme that acetates the hydroxy groups a that lack antibacterial activity,Synthesis of Chloramphenicol i il Bi, a are Brominal oul oni mo i ioe im pattroacetophenone Pritrophenacy| bromide camino pairoacetophenone HCHO (he of “od, ig aan Prien O8 Ptroacetamido acetophenone — oH oH { Ne, fone AYA a Nuninium HCOOH isopropoide On OH ' on Racemic form Deprotection Ohiren of yoxcooc, oy (ami Nh trmefat My wv ‘oH ay VW enantiomerically pure ChloramphenicolClinical applications ¥ thas een reported tobe the drug of choice forthe treatment f typhus and typhoid f ¥ Chloramphenicol is recommended spectfically for the treatment of serious infections strains of Gram-positive and Gram-negative bacteria that have developed resistance to p and ampicilin, such as H. influenzae, Salmonella typhi, $. pneumoniae, B, fags, meningitis, ¥ Because of its penetration into the central nervous system, chloramphenicol is particularly important alternative therapy for meningtis, ¥Chlorampteicl slo ued forthe treatment of rickets nets, suchas Rocky Mourn spotted fever, | ¥ Because its bitter, this antibiotic is administered orally either in capsules or asthe vine Chloramphenicol palmitate insoluble in water and may be suspended in aqueous vehicles fo Liquid dosage forms (either oral or systematic),Lincosamides ar bacteriostatic a ff ganism called Strep The lincosamice antibiotics have simi the same fashion (Inhibit polypeptide efflux channel and agents and was isolated from a solo the clinically useful clindamycin with increased activity ¥ Iti used for osteomyelitis (Done) or joint infection inflammatory disease, strep throat, pneumoria, ac ( some cases of me RSA), be used to treat media (middle ear infection), and endocarditis, ¥ It can also be used to treat acne a resistant Staphylococus aureus (UI ¥ Incombinaton with quinine, i a ¥ is available as oral, systematic as well as topical me ¥ It is very soluble in water, The topically used cli tibacterial properties tothe mac s, pelvic alaia, translocation), Lincomyecin was t omyceslincolnensis, Chemical mo te otitis N thiclin- | HN Ih a Clindamycin ication, amycin BS / Phosphate atac phosphate is a phosphate-ester prodrug of clindamycin, CS? GTN campusSummary Mechanism of protein synthesis inhibition |, acrid and Clindamycin bind to 50s and block polypeptide exit tunnel and interfere with translocation {iia fate jue a“ ~ nda ncn misreading and interferes wath translocation bind to 50s and prevent formation of 30s and 50s X ribosomal complex inhibit binding of tRNA to ribosomal complex Chloramphenicol binds to 50s and inhibits peptide bond J formation X tae X bert1, Compare and contrast the mechanisms of ation ofthe tetracyclines and the macral 2, Notes on SAR and applications of tetracyclines, 3, Write synthesis and mechanism of action of Chloramphenical, 4, Compare and contrast aminoglycosides and macrolides, 5, Write structure and clinical uses of Streptomycin, 6, Describe the degradation products of tetracycline, 7. Structure and uses of Cindamycin, 8, Why can’t streptomycin inhibit mammalian protein synthesis? 9, Name te structure of sugar moieties present in streptomycin and macrolide antibiotics, 10.\ite the structure and mechanism of action of Kanamycin, 11. Cive two examples each fom antibiotics belonging to aminoglycoside and macrlices, 12.Are Chloramphenicol and Clindamycin contraindicated, justi? 13. Differentiate bactericidal and bacteriostatic agents with suitable examples with nile 4 Name a broad-spectrum antibacterial having protein synthetic inhibitory activity along wi a15.\hat is eptetracyclin, now itis formed what sits antibacterial spectrum? {6.Uhy is erythromycin unstable n acid and has to be administered as enteric coated? 17. Adverse effects of tetracycline and ther mechanism {8.Doxycyctne and Minocycline exhibit better absorption than tetracycline, True/False; ju 19, \hich configuration of Chloramphenicol is active? Give stereochemistry. 20.2-Deowstreptamine i integral to which structure ofthe following? [Streptomycin OR Neomycin OR Kanamycin OR Azithromycin} 21 Aminolycoide and Chloramphenicol bind to which ribosomal subunit? 2. Translocation step in bacterial protein synthesis is blocked by which antibacterial cg 23 Which antibacterial drug inhibits peptide bond formation? 24 Tetracycline antibiotics are based on which chemical scaffold? 25, What are the strategies to prevent acid senstvty of erythromycin? /