Volume 16 Number 3 1988 Nucleic Acids Research The nucleotide sequence of the tetracycline resistance determinant tetM from Ureaplasma urealyticum Ray Sanchez-Pescador, Janice T.Brown1, Marilyn Roberts' and Mickey S.Urdea Chiron Corporation, 4560 Horton St., Emeryville, CA 94608 and 'Department of Pathobiology, SC-38, School of Public Health and Community Medicine, University of Washington, Seattle, WA 98195, USA Submitted November 16, 1987 The tetracycline resistance gene, tetM, first detected in Gram-positive streptococci (1), has also been found in pathogenic Gram-negative bacteria (2). This determinant cornfers high-level resistance and is frequently located on conjugative transposable elements (3). Recently, a tetM gerie has been detected in and cloned from Ureaplasma urealyticum (J.T. Brown and M.Roberts,submitted). Here we report its nucleotide sequence. A total of 3425 bp were sequenced by the Sanger method (4) using unpurified synthetic primers (5). An open reading frame (ORF), starting from a Met codon at position 956 that is preceded by a putative ribosome binding site, TGGAGGA, codes for a protein of 639 amino acids (MW= 72,611 Daltons). This molecular weight is in agreement. with previous data (6). The amino acid composition suggests a cytoplasmic protein (6), differing from other tetracycline resistance determinants invoLved in tetracycline efflux (7). Comparison of the tetM nucleotide sequence from Ureaplasma (isolated in Seattle in 1984) with that obtained from S. pneumoniae (6; isolated nearly 7 years earlier in Paris) reveals a close relationship. The ORFs show a 95% homology at the amirno acid level. Most of the differences are conservative. Remarkably then, the tetM gene has been conserved over time, geographical distance and passage between distantly related bacteria (9). It has been suggested that Mycoplasmatales (such as U. uirealyticum) do not utilize the typical TGG codon for Trp, but instead employ the termination triplet TGA (8). The tetM gene described here contains four TGG codons, implying normal Trp codon usage in U. urealyticum. The sequence does not contain a TGA termination; therefore, whether or not. the organism can additionally utilize TGA as a Trp codon remains unclear. 1 GCTCS ACAGAThACAGTAGCAAGATAtSCCt?TATCGTCtC?GTAAGAGAmCC AAG"ATGACCOTAGAGCCAACTGGCGACAATGACGTTAATGTAACCTACAGTGTAGACCAG 121 CTCASTACAGAGGGAGAAAATACAAACACCGTCCACTCTCCTATATACSCAGCTAStGAGATCSSCT CAAAtAS"SACTCCSTAACAACCCACCATtACCAATACCTAAG 2 41 AAAtCAAGTTATAAACCAAAACCCAtSCAAACT .. CACCCTTCATTCCATTACAACCAA2tCAAATCAASCACSTT2SSAACCACOSTCSTCATCCTtASCCTACAOCGACACCCACT 3 61 GAACTSTCCTACTAtCSCGAATCACtDOATAtSAAACCATCCAAAAGAGTACATCT TTCAAGAACTCTAzAATCCTATTCACJLATCGTAACOATAATCAAGCTCACCOTJAATCGCCTCA 481 601 721 641 161 1081 1201 1321 CACSC"iCSASAtC"CCAGCACACCAAACCAACCCACCSATCTCAASSSGASTCCGTACttCAAAACAACOGGACSAAttCCAttsATOAAAATAACAKATAtASCtAACATAtSACA GATACSSWtGAATCASCtACtCtetetCAtAAAzA&AStGAOAtTCCStTACAAKATGtCTCSCACG tCCtASSTTAmAGACtCACtTASAAAAGCAASTAAtAAGCOAAt^ GTA TTCSS"AATAAACtCTCAATTTGATAGCGGGAACAKATASATASSTGSCCe * *...AGGACCCCSAGSTSTSTCSTCCCAGTTTAAGJATSACCTTTATCAITCATSTCTAAACTATCCA GAGAATATCTrTATCCTTTGTATACCSATGGTSATGCAtAAAAAtCCCAGTGATAAAAGtATTSASCAC2CGGASTTSATTrATIACATTGAGGTTAATCGYCAC4AAAG?ACAtGAAA c a G c~~~G h 11-I1 Asal loGyVlLouAlaRiValApAlgaGlyLy&TlartirL-uThrGlu -rL uL uSyrA&SnXrGlyA^latle?rGluL uC1 2 V IAs LyodlyTbrAzr ATTASSJATATTGGCC TTAC.CTCASC.tSCA sAGGAAAAACTACCSTAACAGAAArCCSSATSAtATAACACTGGAGCGASSACAGAAtSAC =aaCAAm SACAACeACC Al& L ThlrAApAsatArLAuLAuGluArgGlmAZelylAlebrzll-TkrGlyllerS rPb ClT aS.TrLsValAslll AsptlrProAlylaNthapPb.L.u ACGGATAATAC4X frSAGAACGSCAGAOACeAATTACAASSCA"CAGAtATACCSCTTSSAT.-AAAASTACGAAGG'tAACAATCACAACACeCAr.GACATAtGOASTTCTA G CG A T SAl lVlyrr$ eSerVal-L u&rlAalLuLeul-eS-rAlaLy AspClyValGI-AlaGlaThrArgll*LeuPll<isAlLoLuArgLyabtClyllePro GCAGAACSATACGTC ASSAtCAGCSSSS TGGACA TCtACTr.A2TSTGCS AAACGAUCSTACAAC.CACAAACTCGTATATST;.TTCASCCACTTACGwAAAATCCGGATTCCC A A T ThzXlePhePbeIlAsaLyzAlAeAs AAlAssG yleAspLeuSTATC rVlT?yrzlAsp-leLyalu.Ly LeuSerAlagluXleVaelIlLysGInLyaValGluLeuTyrpro ACAAtCTSSSSTATCAATAAGAI?GACCAAAATGGAATtGA? AAAACGAtACA AGAAACCtCTGCCCAAAT CSAATCAAACAGAAAGTAGAACTGTAtCCT a C 1216 © R L Press Limited, Oxford, 2 42 82 122 162 1216 C I R L Press Limited, Oxford, England.