{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,9]],"date-time":"2026-10-09T20:06:41Z","timestamp":1791576401732,"version":"4.3.4"},"reference-count":52,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2016,9,22]],"date-time":"2016-09-22T00:00:00Z","timestamp":1474502400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001659","name":"Deutsche Forschungsgemeinschaft","doi-asserted-by":"publisher","award":["HI 291\/12-1"],"award-info":[{"award-number":["HI 291\/12-1"]}],"id":[{"id":"10.13039\/501100001659","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Marie Curie Action Career Integration Grant","award":["PCIG14-GA-2013-632072"],"award-info":[{"award-number":["PCIG14-GA-2013-632072"]}]},{"DOI":"10.13039\/501100003329","name":"Ministerio de Econom\u00eda Y Competitividad","doi-asserted-by":"publisher","award":["CTQ2014-55907-R"],"award-info":[{"award-number":["CTQ2014-55907-R"]}],"id":[{"id":"10.13039\/501100003329","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Antibiotics"],"abstract":"<jats:p>Omadacycline is an aminomethylcycline antibiotic with potent activity against many Gram-positive and Gram-negative pathogens, including strains carrying the major efflux and ribosome protection resistance determinants. This makes it a promising candidate for therapy of severe infectious diseases. Omadacycline inhibits bacterial protein biosynthesis and competes with tetracycline for binding to the ribosome. Its interactions with the 70S ribosome were, therefore, analyzed in great detail and compared with tigecycline and tetracycline. All three antibiotics are inhibited by mutations in the 16S rRNA that mediate resistance to tetracycline in Brachyspira hyodysenteriae, Helicobacter pylori, Mycoplasma hominis, and Propionibacterium acnes. Chemical probing with dimethyl sulfate and Fenton cleavage with iron(II)-complexes of the tetracycline derivatives revealed that each antibiotic interacts in an idiosyncratic manner with the ribosome. X-ray crystallography had previously revealed one primary binding site for tetracycline on the ribosome and up to five secondary sites. All tetracyclines analyzed here interact with the primary site and tetracycline also with two secondary sites. In addition, each derivative displays a unique set of non-specific interactions with the 16S rRNA.<\/jats:p>","DOI":"10.3390\/antibiotics5040032","type":"journal-article","created":{"date-parts":[[2016,9,22]],"date-time":"2016-09-22T09:59:55Z","timestamp":1474538395000},"page":"32","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":32,"title":["The Novel Aminomethylcycline Omadacycline Has High Specificity for the Primary Tetracycline-Binding Site on the Bacterial Ribosome"],"prefix":"10.3390","volume":"5","author":[{"given":"Corina","family":"Heidrich","sequence":"first","affiliation":[{"name":"Microbiology, Department of Biology, Friedrich-Alexander-Universit\u00e4t Erlangen-N\u00fcrnberg, 91058 Erlangen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sanya","family":"Mitova","sequence":"additional","affiliation":[{"name":"Microbiology, Department of Biology, Friedrich-Alexander-Universit\u00e4t Erlangen-N\u00fcrnberg, 91058 Erlangen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andreas","family":"Schedlbauer","sequence":"additional","affiliation":[{"name":"Structural Biology Unit, CIC bioGUNE, 48160 Derio, Bizkaia, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sean","family":"Connell","sequence":"additional","affiliation":[{"name":"Structural Biology Unit, CIC bioGUNE, 48160 Derio, Bizkaia, Spain"},{"name":"IKERBASQUE, Basque Foundation for Science, 48013 Bilbao, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paola","family":"Fucini","sequence":"additional","affiliation":[{"name":"Structural Biology Unit, CIC bioGUNE, 48160 Derio, Bizkaia, Spain"},{"name":"IKERBASQUE, Basque Foundation for Science, 48013 Bilbao, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Judith","family":"Steenbergen","sequence":"additional","affiliation":[{"name":"Paratek Pharmaceuticals Inc., King of Prussia, PA 19406, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7500-5864","authenticated-orcid":false,"given":"Christian","family":"Berens","sequence":"additional","affiliation":[{"name":"Microbiology, Department of Biology, Friedrich-Alexander-Universit\u00e4t Erlangen-N\u00fcrnberg, 91058 Erlangen, Germany"},{"name":"Institute of Molecular Pathogenesis, Friedrich-Loeffler-Institut, 07743 Jena, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2016,9,22]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"2306","DOI":"10.1128\/AAC.35.11.2306","article-title":"Molecular basis of tetracycline action: Identification of analogs whose primary target is not the bacterial ribosome","volume":"35","author":"Rasmussen","year":"1991","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1143","DOI":"10.1016\/S0092-8674(00)00216-6","article-title":"The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit","volume":"103","author":"Brodersen","year":"2000","journal-title":"Cell"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1829","DOI":"10.1093\/emboj\/20.8.1829","article-title":"Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3","volume":"20","author":"Pioletti","year":"2001","journal-title":"EMBO J."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"3812","DOI":"10.1073\/pnas.1216691110","article-title":"Structural basis for potent inhibitory activity of the antibiotic tigecycline during protein synthesis","volume":"110","author":"Jenner","year":"2013","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"2849","DOI":"10.1128\/AAC.04895-14","article-title":"Structural characterization of an alternative mode of tigecycline binding to the bacterial ribosome","volume":"59","author":"Schedlbauer","year":"2015","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1008","DOI":"10.1038\/nsmb831","article-title":"tRNA selection and kinetic proofreading in translation","volume":"11","author":"Blanchard","year":"2004","journal-title":"Nat. Struct. Mol. Biol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"17","DOI":"10.1111\/j.1749-6632.2011.06354.x","article-title":"The history of the tetracyclines","volume":"1241","author":"Nelson","year":"2011","journal-title":"Ann. N. Y. Acad. Sci."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"232","DOI":"10.1128\/MMBR.65.2.232-260.2001","article-title":"Tetracycline antibiotics: Mode of action, applications, molecular biology, and epidemiology of bacterial resistance","volume":"65","author":"Chopra","year":"2001","journal-title":"Microbiol. Mol. Biol. Rev."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"718","DOI":"10.1128\/CMR.00002-11","article-title":"Food animals and antimicrobials: Impacts on human health","volume":"24","author":"Marshall","year":"2011","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"419","DOI":"10.1007\/s00018-009-0172-6","article-title":"The tetracycline resistome","volume":"67","author":"Thaker","year":"2010","journal-title":"Cell Mol. Life Sci."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"738","DOI":"10.1128\/AAC.43.4.738","article-title":"In vitro and in vivo antibacterial activities of a novel glycylcycline, the 9-t-butylglycylamido derivative of minocycline (GAR-936)","volume":"43","author":"Petersen","year":"1999","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"7044","DOI":"10.1128\/AAC.01536-15","article-title":"Structure-activity relationship of the aminomethylcyclines and the discovery of omadacycline","volume":"59","author":"Honeyman","year":"2015","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1279","DOI":"10.1128\/AAC.01066-13","article-title":"Mechanism of action of the novel aminomethylcycline antibiotic omadacycline","volume":"58","author":"Draper","year":"2014","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1127","DOI":"10.1128\/AAC.01242-13","article-title":"In vitro and in vivo antibacterial activities of omadacycline, a novel aminomethylcycline","volume":"58","author":"Macone","year":"2014","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"592","DOI":"10.1093\/jac\/dkh125","article-title":"Comparison of tetracycline and tigecycline binding to ribosomes mapped by dimethylsulphate and drug-directed Fe2+ cleavage of 16S rRNA","volume":"53","author":"Bauer","year":"2004","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"11829","DOI":"10.1021\/bi0506066","article-title":"Tigecycline is modified by the flavin-dependent monooxygenase TetX","volume":"44","author":"Moore","year":"2005","journal-title":"Biochemistry"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"2156","DOI":"10.1128\/AAC.01499-05","article-title":"Functional, biophysical, and structural bases for antibacterial activity of tigecycline","volume":"50","author":"Olson","year":"2006","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Nelson, M., Hillen, W., and Greenwald, R.A. (2002). Tetracyclines in Biology, Chemistry and Medicine, Birkh\u00e4user.","DOI":"10.1007\/978-3-0348-8306-1"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"5650","DOI":"10.1128\/AAC.00948-12","article-title":"A randomized, evaluator-blind, phase 2 study comparing the safety and efficacy of omadacycline to those of linezolid for treatment of complicated skin and skin structure infections","volume":"56","author":"Noel","year":"2012","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1702","DOI":"10.1128\/AAC.42.7.1702","article-title":"16S rRNA mutation associated with tetracycline resistance in a Gram-positive bacterium","volume":"42","author":"Ross","year":"1998","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"3708","DOI":"10.1128\/JB.187.11.3708-3712.2005","article-title":"16S rRNA mutations that confer tetracycline resistance in Helicobacter pylori decrease drug binding in Escherichia coli ribosomes","volume":"187","author":"Nonaka","year":"2005","journal-title":"J. Bacteriol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"245","DOI":"10.1016\/j.vetmic.2007.02.019","article-title":"Decreased susceptibility to doxycycline associated with a 16S rRNA gene mutation in Brachyspira hyodysenteriae","volume":"123","author":"Pringle","year":"2007","journal-title":"Vet. Microbiol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1390","DOI":"10.1093\/jac\/dkn118","article-title":"Reduced susceptibility to tetracyclines is associated in vitro with the presence of 16S rRNA mutations in Mycoplasma hominis and Mycoplasma pneumoniae","volume":"61","author":"Renaudin","year":"2008","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"796","DOI":"10.1128\/AAC.03876-14","article-title":"16S rRNA gene mutations associated with decreased susceptibility to tetracycline in Mycoplasma bovis","volume":"59","author":"Amram","year":"2015","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"389","DOI":"10.1038\/327389a0","article-title":"Interaction of antibiotics with functional sites in 16S ribosomal RNA","volume":"327","author":"Moazed","year":"1987","journal-title":"Nature"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1971","DOI":"10.1073\/pnas.96.5.1971","article-title":"An Escherichia coli strain with all chromosomal rRNA operons inactivated: Complete exchange of rRNA genes between bacteria","volume":"96","author":"Asai","year":"1999","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"112","DOI":"10.1016\/0147-619X(81)90058-5","article-title":"Construction and fine mapping of recombinant plasmids containing the rrnB ribosomal RNA operon of E. coli","volume":"6","author":"Brosius","year":"1981","journal-title":"Plasmid"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1097","DOI":"10.1007\/s00775-008-0395-2","article-title":"Specific binding of divalent metal ions to tetracycline and to the Tet repressor\/tetracycline complex","volume":"13","author":"Palm","year":"2008","journal-title":"J. Biol. Inorg. Chem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"22","DOI":"10.1021\/bi00001a004","article-title":"Proximity mapping of the Tet repressor-tetracycline-Fe2+ complex by hydrogen peroxide mediated protein cleavage","volume":"34","author":"Ettner","year":"1995","journal-title":"Biochemistry"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"5113","DOI":"10.1128\/JB.184.18.5113-5120.2002","article-title":"Fe2+-tetracycline-mediated cleavage of the Tn10 tetracycline efflux protein TetA reveals a substrate binding site near glutamine 225 in transmembrane helix 7","volume":"184","author":"McMurry","year":"2002","journal-title":"J. Bacteriol."},{"key":"ref_31","unstructured":"Westhof, E., Bindereif, A., Sch\u00f6n, A., and Hartmann, R.K. (2014). Handbook of RNA Biochemistry, WILEY-VCH. [2nd ed.]."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"175","DOI":"10.1046\/j.0014-2956.2001.02636.x","article-title":"Inhibition of nuclear pre-mRNA splicing by antibiotics in vitro","volume":"269","author":"Hertweck","year":"2002","journal-title":"Eur. J. Biochem."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1219","DOI":"10.1093\/nar\/25.6.1219","article-title":"Interaction of tetracycline with RNA: Photoincorporation into ribosomal RNA of Escherichia coli","volume":"25","author":"Oehler","year":"1997","journal-title":"Nucleic Acids Res."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"985","DOI":"10.1016\/0092-8674(86)90813-5","article-title":"Transfer RNA shields specific nucleotides in 16S ribosomal RNA from attack by chemical probes","volume":"47","author":"Moazed","year":"1986","journal-title":"Cell"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"16576","DOI":"10.1074\/jbc.274.23.16576","article-title":"Effects of tetracycline and spectinomycin on the tertiary structure of ribosomal RNA in the Escherichia coli 30S ribosomal subunit","volume":"274","author":"Noah","year":"1999","journal-title":"J. Biol. Chem."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"309","DOI":"10.1016\/0022-2836(88)90243-4","article-title":"Probing the assembly of the 3\u2019 major domain of 16 S ribosomal RNA. Quaternary interactions involving ribosomal proteins S7, S9 and S19","volume":"200","author":"Powers","year":"1988","journal-title":"J. Mol. Biol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"2131","DOI":"10.1128\/JB.184.8.2131-2140.2002","article-title":"Mutations in the 16S rRNA genes of Helicobacter pylori mediate resistance to tetracycline","volume":"184","author":"Trieber","year":"2002","journal-title":"J. Bacteriol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"277","DOI":"10.1089\/mdr.2000.6.277","article-title":"Mutations in the interdomain loop region of the tetA(A) tetracycline resistance gene increase efflux of minocycline and glycylcyclines","volume":"6","author":"Tuckman","year":"2000","journal-title":"Microb. Drug Resist."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"2065","DOI":"10.1128\/AAC.01198-06","article-title":"Tigecycline efflux as a mechanism for nonsusceptibility in Acinetobacter baumannii","volume":"51","author":"Peleg","year":"2007","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"133","DOI":"10.1016\/j.ijantimicag.2013.04.017","article-title":"The tetA gene decreases tigecycline sensitivity of Salmonella enterica isolates","volume":"42","author":"Akiyama","year":"2013","journal-title":"Int. J. Antimicrob. Agents"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"297","DOI":"10.1128\/AAC.01727-13","article-title":"Molecular epidemiology and mechanisms of tigecycline resistance in clinical isolates of Acinetobacter baumannii from a chinese university hospital","volume":"58","author":"Deng","year":"2014","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"1707","DOI":"10.1128\/AAC.01803-13","article-title":"Genomics of KPC-producing Klebsiella pneumoniae sequence type 512 clone highlights the role of RamR and ribosomal S10 protein mutations in conferring tigecycline resistance","volume":"58","author":"Villa","year":"2014","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"193","DOI":"10.1016\/0162-0134(88)80049-7","article-title":"Metal ion-tetracycline interactions in biological fluids. Part 8. Potentiometric and spectroscopic studies on the formation of Ca(II) and Mg(II) complexes with 4-dedimethylamino-tetracycline and 6-desoxy-6-demethyl-tetracycline","volume":"33","author":"Lambs","year":"1988","journal-title":"J. Inorg. Biochem."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"3502","DOI":"10.1093\/nar\/22.17.3502","article-title":"Collection of small subunit (16S- and 16S-like) ribosomal RNA structures: 1994","volume":"22","author":"Gutell","year":"1994","journal-title":"Nucleic Acids Res."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"2594","DOI":"10.1093\/nar\/gkh583","article-title":"Mapping of the second tetracycline binding site on the ribosomal small subunit of E. coli","volume":"32","author":"Anokhina","year":"2004","journal-title":"Nucleic Acids Res."},{"key":"ref_46","first-page":"9","article-title":"DH5\u03b1 competent cells","volume":"8","author":"Hanahan","year":"1986","journal-title":"Focus"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"4290","DOI":"10.1073\/pnas.81.14.4290","article-title":"Ribonuclease T: New exoribonuclease possibly involved in end-turnover of tRNA","volume":"81","author":"Deutscher","year":"1984","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"14077","DOI":"10.1074\/jbc.273.23.14077","article-title":"The vacb gene required for virulence in Shigella flexneri and Escherichia coli encodes the exoribonuclease RNase R","volume":"273","author":"Cheng","year":"1998","journal-title":"J. Biol. Chem."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"725","DOI":"10.1006\/jmbi.2001.5359","article-title":"Crystal structure of the 30S ribosomal subunit from Thermus thermophilus: Structure of the proteins and their interactions with 16S RNA","volume":"316","author":"Brodersen","year":"2002","journal-title":"J. Mol. Biol."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"292","DOI":"10.1016\/S0076-6879(00)17021-1","article-title":"Preparation of functional ribosomal complexes and effect of buffer conditions on tRNA positions observed by cryoelectron microscopy","volume":"317","author":"Blaha","year":"2000","journal-title":"Methods Enzymol."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"163","DOI":"10.1016\/S1074-5521(98)90061-8","article-title":"Visualizing metal-ion-binding sites in group I introns by iron(II)-mediated Fenton reactions","volume":"5","author":"Berens","year":"1998","journal-title":"Chem. Biol."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"399","DOI":"10.1016\/0022-2836(86)90441-9","article-title":"Rapid chemical probing of conformation in 16S ribosomal RNA and 30S ribosomal subunits using primer extension","volume":"187","author":"Moazed","year":"1986","journal-title":"J. Mol. Biol."}],"container-title":["Antibiotics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2079-6382\/5\/4\/32\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T19:31:35Z","timestamp":1760211095000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2079-6382\/5\/4\/32"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2016,9,22]]},"references-count":52,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2016,12]]}},"alternative-id":["antibiotics5040032"],"URL":"https:\/\/doi.org\/10.3390\/antibiotics5040032","relation":{},"ISSN":["2079-6382"],"issn-type":[{"value":"2079-6382","type":"electronic"}],"subject":[],"published":{"date-parts":[[2016,9,22]]}}}