The Effects of Tiamulin, a Semisynthetic Pleuromutilin Derivative, on Bacterial Polypeptide Chain Initiation

Abstract

Tiamulin, a water-soluble and highly effective semisynthetic derivative of pleuromutilin leads to the formation of physiologically inactive polypeptide chain initiation complexes which readily decompose and do not enter the phase of peptide chain elongation. Once elongation has begun it continues even in the presence of tiamulin as has been shown by measuring the formation of N-acetylphenylalanine-poly(phenyla1anine). The formation of abortive initiation complexes was observed regardless of whether AcPhe-tRNA of Met-tRNA was used as an initiator or whether artificial messengers or a natural messenger, like R17 bacteriophage RNA, was used. When this drug was acting on whole cells, it led to the disappearance of polysomes. The only structures which could be detected were of the monosome size. Therefore, polysomes seem to elongate the polypeptide chains in whole cells in the presence of this antibiotic, but since effective reinitiation is blocked, the polysome pool of the cell soon becomes depleted.Pleuromutilin, a tricyclic diterpene antibiotic which is produced by a basidiomycete, is active against bacteria and mycoplasms. When the glycolic acid residue of the natural product is replaced by basic thio-ether side chains the antibacterial activity is increased considerably. One semisynthetic compound with these chemical characteristics, the 14-deoxy-14-[(2-diethylaminoethy1)mercapto-acetoxy]mutilin hydrogen fumarate, has been given the generic name 'tiamulin'. The formulae of both the natural substance and the semisynthetic compound were described in previous communications from this institute [l, 21. In order to elucidate the mode of action of pleuromutilin and its derivatives, tiamulin and chemically closely related substances have been selected as model compounds. By experiments described in an earlier communication [2] these pleuromutilin derivatives were characterized as inhibitors of prokaryotic ribosomes. At micromolar concentrations they cause a dramatic decline in the poly(pheny1alanine) synthesis, the synthesis of formylmethionine-puromycin or that of the initial dipeptide of the bacteriophage R17 coat protein. In the latter reaction it appeared as if the formation of initiation complexes was only slightly affected, while the binding of alanyl-tRNA to the ribosomal A-site was inhibited to a large extent. Elongation of nascent polypeptides on natural polysomes, when assayed in vitro, showed little sensitivity towards tiamulin. The formation of the first peptide bond, involving the initiating formylmethionine residue, thus appears to be particularly sensitive towards pleuromutilin derivatives.In order to explain this sensitivity of the first peptide bond, the present work was undertaken. The result showed that although initiation complexes are formed in the presence of the drug, they are defective and therefore do not allow further propagation of the protein chain.Abbreviations. AcPhe-poly(Phe), N-acetylphenylalanine-poly-(phenylalanine); EF-T, mixture of elongation factors EF-Tu and EF-Ts; Hepes, 4-(2-hy...

Induced-fit tightens pleuromutilins binding to ribosomes and remote interactions enable their selectivity

“…1), undergo reduced metabolism (16), thus more suitable for oral administration. As observed biochemically, pleuromutilins interfere with peptide bond formation (17)(18)(19)(20)(21). Consistently, the only available crystal structure of a pleuromutilin antibiotic, tiamulin, bound to the large ribosomal subunit verifies binding to the PTC (22).…”

Ribosomal binding region for the antibiotic tiamulin: stoichiometry, subunit location, and affinity for various analogs

“…1), acts as an inhibitor of bacterial protein biosynthesis (1,4). The 19,20-dihydro derivative has been shown to bind strongly to ribosomes from Escherichia coli strain D10.…”

Effect of antibiotics on the bacterial population of the rabbit caecum

“…This observation is not particularly surprising; however, the combination of data from Table 3 and Fig. Chlortetracycline is known to inhibit protein synthesis by preventing binding of aminoacyl-tRNA to ribosomes, tiamulin inhibits protein synthesis by specifically targeting the large subunit of the bacterial ribosome and inhibiting peptide bond formation, and bacitracin inhibits cell wall synthesis (Davis & Feingold, 1962;Dornhelm & Hogenauer, 1978;Chopra & Roberts, 2001). Chlortetracycline is known to inhibit protein synthesis by preventing binding of aminoacyl-tRNA to ribosomes, tiamulin inhibits protein synthesis by specifically targeting the large subunit of the bacterial ribosome and inhibiting peptide bond formation, and bacitracin inhibits cell wall synthesis (Davis & Feingold, 1962;Dornhelm & Hogenauer, 1978;Chopra & Roberts, 2001).…”