{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,24]],"date-time":"2026-09-24T23:10:01Z","timestamp":1790291401660,"version":"4.1.0"},"reference-count":48,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2021,6,28]],"date-time":"2021-06-28T00:00:00Z","timestamp":1624838400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100010751","name":"Fondo Nacional de Desarrollo Cient\u00edfico, Tecnol\u00f3gico y de Innovaci\u00f3n Tecnol\u00f3gica","doi-asserted-by":"publisher","award":["154-2017-FONDECYT"],"award-info":[{"award-number":["154-2017-FONDECYT"]}],"id":[{"id":"10.13039\/501100010751","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100010751","name":"Fondo Nacional de Desarrollo Cient\u00edfico, Tecnol\u00f3gico y de Innovaci\u00f3n Tecnol\u00f3gica","doi-asserted-by":"publisher","award":["036-2019-FONDECYT-BM-INC-INV"],"award-info":[{"award-number":["036-2019-FONDECYT-BM-INC-INV"]}],"id":[{"id":"10.13039\/501100010751","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100010751","name":"Fondo Nacional de Desarrollo Cient\u00edfico, Tecnol\u00f3gico y de Innovaci\u00f3n Tecnol\u00f3gica","doi-asserted-by":"publisher","award":["406-2019-FONDECYT"],"award-info":[{"award-number":["406-2019-FONDECYT"]}],"id":[{"id":"10.13039\/501100010751","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100010750","name":"Inn\u00f3vate Per\u00fa","doi-asserted-by":"publisher","award":["297-InnovatePeru-2016"],"award-info":[{"award-number":["297-InnovatePeru-2016"]}],"id":[{"id":"10.13039\/501100010750","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Microbiol."],"abstract":"<jats:p>Tetracycline has positively impacted human health as well as the farming and animal industries. Its extensive usage and versatility led to the spread of resistance mechanisms followed by the development of new variants of the antibiotic. Tetracyclines inhibit bacterial growth by impeding the binding of elongator tRNAs to the ribosome. However, a small number of reports indicated that Tetracyclines could also inhibit translation initiation, yet the molecular mechanism remained unknown. Here, we use biochemical and computational methods to study how Oxytetracycline (Otc), Demeclocycline (Dem), and Tigecycline (Tig) affect the translation initiation phase of protein synthesis. Our results show that all three Tetracyclines induce Initiation Factor IF3 to adopt a compact conformation on the 30S ribosomal subunit, similar to that induced by Initiation Factor IF1. This compaction was faster for Tig than Dem or Otc. Furthermore, all three tested tetracyclines affected IF1-bound 30S complexes. The dissociation rate constant of IF1 in early 30S complexes was 14-fold slower for Tig than Dem or Otc. Late 30S initiation complexes (30S pre-IC or IC) exhibited greater IF1 stabilization by Tig than for Dem and Otc. Tig and Otc delayed 50S joining to 30S initiation complexes (30S ICs). Remarkably, the presence of Tig considerably slowed the progression to translation elongation and retained IF1 in the resulting 70S initiation complex (70S IC). Molecular modeling of Tetracyclines bound to the 30S pre-IC and 30S IC indicated that the antibiotics binding site topography fluctuates along the initiation pathway. Mainly, 30S complexes show potential contacts between Dem or Tig with IF1, providing a structural rationale for the enhanced affinity of the antibiotics in the presence of the factor. Altogether, our data indicate that Tetracyclines inhibit translation initiation by allosterically perturbing the IF3 layout on the 30S, retaining IF1 during 70S IC formation, and slowing the transition toward translation elongation. Thus, this study describes a new complementary mechanism by which Tetracyclines may inhibit bacterial protein synthesis.<\/jats:p>","DOI":"10.3389\/fmicb.2021.682682","type":"journal-article","created":{"date-parts":[[2021,6,28]],"date-time":"2021-06-28T01:46:01Z","timestamp":1624844761000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":42,"title":["A Complementary Mechanism of Bacterial mRNA Translation Inhibition by Tetracyclines"],"prefix":"10.3389","volume":"12","author":[{"given":"Victor","family":"Barrenechea","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maryhory","family":"Vargas-Reyes","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Miguel","family":"Quiliano","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pohl","family":"Mil\u00f3n","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2021,6,28]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"20530","DOI":"10.1073\/pnas.2008671117","article-title":"Sarecycline interferes with tRNA accommodation and tethers mRNA to the 70S ribosome.","volume":"117","author":"Batool","year":"2020","journal-title":"Proc. Natl. Acad. Sci. U.S.A."},{"key":"B2","doi-asserted-by":"publisher","first-page":"4056","DOI":"10.1002\/j.1460-2075.1995.tb00077.x","article-title":"X-ray crystallography shows that translational initiation factor IF3 consists of two compact alpha\/beta domains linked by an alpha-helix.","volume":"14","author":"Biou","year":"1995","journal-title":"EMBO J."},{"key":"B3","doi-asserted-by":"publisher","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":"B4","doi-asserted-by":"publisher","first-page":"498","DOI":"10.1126\/science.1057766","article-title":"Crystal structure of an initiation factor bound to the 30S ribosomal subunit.","volume":"291","author":"Carter","year":"2001","journal-title":"Science"},{"key":"B5","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2014.00334","article-title":"Use of antibiotics as feed additives: a burning question.","volume":"5","author":"Chattopadhyay","year":"2014","journal-title":"Front. Microbiol."},{"key":"B6","doi-asserted-by":"publisher","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. R."},{"key":"B7","doi-asserted-by":"publisher","DOI":"10.3390\/antibiotics5040038","article-title":"Conformational Response of 30S-bound IF3 to A-Site Binders Streptomycin and Kanamycin.","volume":"5","author":"Chulluncuy","year":"2016","journal-title":"Antibiotics"},{"key":"B8","doi-asserted-by":"publisher","first-page":"98","DOI":"10.1016\/0006-291x(65)90954-x","article-title":"Binding of tetracycline to the 30S ribosomes and to polyuridylic acid.","volume":"20","author":"Connamacher","year":"1965","journal-title":"Biochem. Biophys. Res. Co."},{"key":"B9","doi-asserted-by":"publisher","DOI":"10.1097\/00007691-198206000-00001","article-title":"Doxycycline.","volume":"4","author":"Cunha","year":"1982","journal-title":"Ther. Drug Monit."},{"key":"B10","doi-asserted-by":"publisher","first-page":"462","DOI":"10.1086\/367622","article-title":"Tetracycline therapy: update.","volume":"36","author":"Eliopoulos","year":"2003","journal-title":"Clin. Infect. Dis."},{"key":"B11","doi-asserted-by":"publisher","first-page":"628","DOI":"10.1038\/nsmb.2554","article-title":"Conformational selection of translation initiation factor 3 signals proper substrate selection.","volume":"20","author":"Elvekrog","year":"2013","journal-title":"Nat. Struct. Mol. Biol."},{"key":"B12","doi-asserted-by":"publisher","first-page":"285","DOI":"10.1016\/j.molcel.2006.12.011","article-title":"The real-time path of translation factor IF3 onto and off the ribosome.","volume":"25","author":"Fabbretti","year":"2007","journal-title":"Mol. Cell."},{"key":"B13","doi-asserted-by":"publisher","first-page":"449","DOI":"10.1042\/bj0870449","article-title":"The inhibition of incorporation of leucine into protein of cell-free systems from rat liver and Escherichia coli by chlortetracycline.","volume":"87","author":"Franklin","year":"1963","journal-title":"Biochem. J."},{"key":"B14","doi-asserted-by":"publisher","first-page":"2080","DOI":"10.2174\/1568026618666181025092546","article-title":"Antibiotics targeting the 30S ribosomal subunit: a lesson from nature to find and develop new drugs.","volume":"18","author":"Giuliodori","year":"2019","journal-title":"Curr. Top. Med. Chem."},{"key":"B15","doi-asserted-by":"publisher","first-page":"10700","DOI":"10.1093\/nar\/gkv869","article-title":"Directional transition from initiation to elongation in bacterial translation.","volume":"43","author":"Goyal","year":"2015","journal-title":"Nucleic Acids Res."},{"key":"B16","doi-asserted-by":"publisher","first-page":"80","DOI":"10.1002\/ijch.201000006","article-title":"Role of the initiation factors in mRNA start site selection and fMet-tRNA recruitment by bacterial ribosomes.","volume":"50","author":"Gualerzi","year":"2010","journal-title":"Israel J. Chem."},{"key":"B17","doi-asserted-by":"publisher","first-page":"4341","DOI":"10.1007\/s00018-015-2010-3","article-title":"Initiation of mRNA translation in bacteria: structural and dynamic aspects.","volume":"72","author":"Gualerzi","year":"2015","journal-title":"Cell Mol. Life Sci."},{"key":"B18","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2014.00288","article-title":"Benefits and risks of antimicrobial use in food-producing animals.","volume":"5","author":"Hao","year":"2014","journal-title":"Front. Microbiol."},{"key":"B19","doi-asserted-by":"publisher","first-page":"2179","DOI":"10.1128\/aac.48.6.2179-2184.2004","article-title":"Effects of efflux transporter genes on susceptibility of Escherichia coli to tigecycline (GAR-936).","volume":"48","author":"Hirata","year":"2004","journal-title":"Antimicrob. Agents Chemother."},{"key":"B20","doi-asserted-by":"publisher","first-page":"133.e13","DOI":"10.1016\/j.cell.2016.08.074","article-title":"Large-scale movements of IF3 and tRNA during bacterial translation initiation.","volume":"167","author":"Hussain","year":"2016","journal-title":"Cell"},{"key":"B21","doi-asserted-by":"publisher","first-page":"321","DOI":"10.1128\/iai.1.4.321-326.1970","article-title":"Activity of minocycline against R factor-carrying enterobacteriaceae1.","volume":"1","author":"Jarolmen","year":"1970","journal-title":"Infect. Immun."},{"key":"B22","doi-asserted-by":"publisher","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. U.S.A."},{"key":"B23","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pbio.1001095","article-title":"The Cryo-EM structure of a complete 30S translation initiation complex from Escherichia coli.","volume":"9","author":"Juli\u00e1n","year":"2011","journal-title":"PLoS Biol."},{"key":"B24","doi-asserted-by":"publisher","first-page":"641","DOI":"10.1007\/978-3-540-69375-8_84","author":"Kligman","year":"2014","journal-title":"Pathogenesis and Treatment of Acne and Rosacea."},{"key":"B25","doi-asserted-by":"publisher","first-page":"137","DOI":"10.1016\/0006-291x(64)90243-8","article-title":"Inhibition by tetracyclines of polyuridylic acid directed phenylalanine incorporation in Escherichia coli cell-free systems.","volume":"14","author":"Laskin","year":"1964","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"B26","doi-asserted-by":"publisher","first-page":"611","DOI":"10.1093\/jac\/dki291","article-title":"Tigecycline: what is it, and where should it be used?","volume":"56","author":"Livermore","year":"2005","journal-title":"J. Antimicrob. Chemother."},{"key":"B27","doi-asserted-by":"publisher","first-page":"2179","DOI":"10.1093\/nar\/gkw1251","article-title":"Structure of a 30S pre-initiation complex stalled by GE81112 reveals structural parallels in bacterial and eukaryotic protein synthesis initiation pathways.","volume":"45","author":"L\u00f3pez-Alonso","year":"2017","journal-title":"Nucleic Acids Res."},{"key":"B28","doi-asserted-by":"publisher","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":"B29","doi-asserted-by":"publisher","first-page":"79","DOI":"10.17221\/5681-vetmed","article-title":"Tetracyclines in veterinary medicine and bacterial resistance to them.","volume":"49","author":"Michalova","year":"2012","journal-title":"Vet. Med."},{"key":"B30","doi-asserted-by":"publisher","first-page":"712","DOI":"10.1016\/j.molcel.2008.04.014","article-title":"Kinetic checkpoint at a late step in translation initiation.","volume":"30","author":"Milon","year":"2008","journal-title":"Mol. Cell."},{"key":"B31","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1016\/s0076-6879(07)30001-3","article-title":"Transient kinetics, fluorescence, and FRET in studies of initiation of translation in bacteria.","volume":"430","author":"Milon","year":"2007","journal-title":"Methods Enzymol."},{"key":"B32","doi-asserted-by":"publisher","first-page":"609","DOI":"10.1038\/nsmb.2285","article-title":"Real-time assembly landscape of bacterial 30S translation initiation complex.","volume":"19","author":"Milon","year":"2012","journal-title":"Nat. Struct. Mol. Biol."},{"key":"B33","doi-asserted-by":"publisher","first-page":"1013","DOI":"10.1038\/2171013a0","article-title":"Binding of Formylmethionyl-tRNA to 30S Ribosomal Sub-units.","volume":"217","author":"Mukundan","year":"1968","journal-title":"Nature"},{"key":"B34","doi-asserted-by":"publisher","first-page":"193","DOI":"10.1093\/dnares\/dsw008","article-title":"Comprehensive identification of translation start sites by tetracycline-inhibited ribosome profiling.","volume":"23","author":"Nakahigashi","year":"2016","journal-title":"DNA Res."},{"key":"B35","doi-asserted-by":"publisher","DOI":"10.1101\/579326v1","article-title":"The complete, functional and dynamic cycle of the bacterial Initiation Factor 3.","author":"Nakamoto","year":"2019","journal-title":"BioRxiv"},{"key":"B36","doi-asserted-by":"publisher","first-page":"559","DOI":"10.1515\/hsz-2013-0292","article-title":"Tetracycline antibiotics and resistance mechanisms.","volume":"395","author":"Nguyen","year":"2014","journal-title":"Biol. Chem."},{"key":"B37","doi-asserted-by":"publisher","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":"B38","doi-asserted-by":"publisher","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":"B39","doi-asserted-by":"publisher","first-page":"603","DOI":"10.1111\/j.1432-1033.1976.tb10726.x","article-title":"Specificity and properties of the destabilization, induced by initiation factor IF-3, of ternary complexes of the 30-S ribosomal subunit, aminoacyl-tRNA and polynucleotides.","volume":"67","author":"Risuleo","year":"1976","journal-title":"Eur. J. Biochem."},{"key":"B40","doi-asserted-by":"publisher","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":"B41","doi-asserted-by":"publisher","first-page":"82","DOI":"10.1017\/s1355838299981487","article-title":"Identification of the ribosome binding sites of translation initiation factor IF3 by multidimensional heteronuclear NMR spectroscopy.","volume":"5","author":"Sette","year":"1999","journal-title":"RNA"},{"key":"B42","doi-asserted-by":"publisher","first-page":"416","DOI":"10.1038\/nature07192","article-title":"Structure of the 30S translation initiation complex.","volume":"455","author":"Simonetti","year":"2008","journal-title":"Nature"},{"key":"B43","doi-asserted-by":"publisher","first-page":"965","DOI":"10.1345\/aph.1h543","article-title":"Tigecycline: a novel broad-spectrum antimicrobial.","volume":"41","author":"Slover","year":"2007","journal-title":"Ann. Pharmacother."},{"key":"B44","doi-asserted-by":"publisher","first-page":"1483","DOI":"10.1073\/pnas.55.6.1483","article-title":"Binding of specific sRNA to 30S ribosomal subunits: effect of 50S ribosomal subunits.","volume":"55","author":"Suzuka","year":"1966","journal-title":"Proc. Natl. Acad. Sci. U.S.A."},{"key":"B45","doi-asserted-by":"crossref","first-page":"6807","DOI":"10.1002\/chem.201203502","article-title":"Direct monitoring of initiation factor dynamics through formation of 30S and 70S translation\u2013initiation complexes on a quartz crystal microbalance.","volume":"19","author":"Takahashi","year":"2013","journal-title":"Chem. Eur. J."},{"key":"B46","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pbio.3000593","article-title":"How the initiating ribosome copes with ppGpp to translate mRNAs.","volume":"18","author":"Vinogradova","year":"2020","journal-title":"PLoS Biol."},{"key":"B47","doi-asserted-by":"publisher","first-page":"499","DOI":"10.1016\/0022-2836(75)90242-9","article-title":"Kinetics of the reversible association of ribosomal subunits: stopped-flow studies of the rate law and of the effect of Mg2+.","volume":"93","author":"Wishnia","year":"1975","journal-title":"J. Mol. Biol."},{"key":"B48","doi-asserted-by":"publisher","first-page":"4055","DOI":"10.1039\/c0cc05111j","article-title":"Molecular mechanisms of antibiotic resistance.","volume":"47","author":"Wright","year":"2011","journal-title":"Chem. Commun."}],"container-title":["Frontiers in Microbiology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmicb.2021.682682\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,6,28]],"date-time":"2021-06-28T01:46:04Z","timestamp":1624844764000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmicb.2021.682682\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,6,28]]},"references-count":48,"alternative-id":["10.3389\/fmicb.2021.682682"],"URL":"https:\/\/doi.org\/10.3389\/fmicb.2021.682682","relation":{},"ISSN":["1664-302X"],"issn-type":[{"value":"1664-302X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,6,28]]},"article-number":"682682"}}