Primary structure of an aminoglycoside 6'-N-acetyltransferase AAC(6')-4, fused in vivo with the signal peptide of the Tn3-encoded beta-lactamase.
The gene aacA4 encoding an aminoglycoside 6'-N-acetyltransferase, AAC(6')-4, was cloned from a natural multiresistance plasmid, and its nucleotide sequence was determined. The gene was 600 base pairs (bp) long, and the AAC(6')-4 had a calculated molecular size of 22.4 kilodaltons and an isoelectric point of 5.35. The sequence of the 17 N-terminal amino acids was determined from the purified enzyme. The AAC(6')-4 gene was part of a resistance gene cluster, and its expression was under the control of the regulatory sequences of the beta-lactamase encoded by Tn3. The five N-terminal amino acids were identical to those of the signal peptide of the Tn3-encoded beta-lactamase, and the entire 5' region of aacA4, as far as it was sequenced (354 bp, including the promoter and the ribosome-binding site sequences), was identical to that of the beta-lactamase gene. This led us to presume an in vivo fusion between the beta-lactamase and the acetyltransferase genes. The latter was followed, in a polycistronic arrangement, by an aminoglycoside 3",9-adenylyltransferase gene, aadA, with an intergenic region of 68 bp. At a distance of ca. 1.3 kilobases in the 3' direction, we found remnants of a second Tn3-like element specifying an active beta-lactamase. At their 5' extremities, the two incomplete copies of Tn3, which were in tandem orientation, were interrupted within the resolvase gene. We speculate that Tn3-related sequences have played a role in the process of selection and dissemination of the AAC(6')-4 gene, which specifies resistance to amikacin and related aminoglycosides.
Tran van Nhieu
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Laboratoire de Microbiologie Médicale, Université Paris, France.
Collatz
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D000123:Acetyltransferases / Q000032:analysis / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics; D004352:Drug Resistance, Microbial; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D021382:Protein Sorting Signals / Q000235:genetics*; D011815:R Factors; D012045:Regulatory Sequences, Nucleic Acid; D012689:Sequence Homology, Nucleic Acid; D001618:beta-Lactamases / Q000235:genetics*
D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D021382:Protein Sorting Signals; D000123:Acetyltransferases; C019681:aminoglycoside acetyltransferase; D004262:DNA Restriction Enzymes; D001618:beta-Lactamases