Genetic and biochemical characterization of TRU-1, the endogenous class C β-lactamase from Aeromonas enteropelogenes

Aeromonas; Aeromonas enteropelogenes; Aeromonas hydrophila; Aeromonas punctata; Aeromonas salmonicida; Escherichia coli; Amino Acid Sequence; Base Sequence; Cloning, Molecular; DNA Primers; DNA, Bacterial; Drug Resistance, Bacterial; Genes, Bacterial; Humans; Kinetics; Molecular Sequence Data; Phylogeny; Sequence Homology, Amino Acid; Sequence Homology, Nucleic Acid; Transformation, Genetic

Abstract :

[en] Aeromonas enteropelogenes (formerly A. tructi) was described to be an ampicillin-susceptible and cephalothinresistant Aeromonas species, which suggests the production of a cephalosporinase. Strain ATCC 49803 was susceptible to amoxicillin, cefotaxime, and imipenem but resistant to cefazolin (MICs of 2, 0.032, 0.125, and >256 μg/ml, respectively) and produced an inducible β-lactamase. Cefotaxime-resistant mutants (MIC, 32 μg/ml) that showed constitutive β-lactamase production could be selected in vitro. The gene coding for the cephalosporinase of A. enteropelogenes ATCC 49803 was cloned, and its biochemical properties were investigated. Escherichia coli transformants showing resistance to various β-lactams carried a 3.5-kb plasmid insert whose sequence revealed a 1,146-bp open reading frame (ORF) encoding a class C β-lactamase, named TRU-1, showing the highest identity scores with A. punctata CAV-1 (75%), A. salmonicida AmpC (75%), and A. hydrophila CepH (71%). The blaTRU-1 locus includes open reading frames (ORFs) showing significant homology with genes found in the genomes of other Aeromonas species, although it exhibits a different organization, as reflected by the presence of additional ORFs located downstream of the β-lactamase gene in the A. hydrophila and A. salmonicida genomes. Specific PCR assays were negative for cphA-like and blaOXA-12-like genes in three A. enteropelogenes ATCC strains. Purified TRU-1 showed a broad substrate profile, efficiently hydrolyzing benzylpenicillin, cephalothin, cefoxitin, and, although with significantly lower turnover rates, oxyiminocephalosporins. Cephaloridine and cefepime were poorly recognized by the enzyme, as reflected by the high K m values observed with these substrates. Thus far, A. enteropelogenes represents the only known example of an Aeromonas species that produces only one β-lactamase belonging to molecular class C. Copyright © 2010, American Society for Microbiology. All Rights Reserved.

Disciplines :

Biochemistry, biophysics & molecular biology
Microbiology

Author, co-author :

De Luca, F.; Dipartimento di Biologia Molecolare, Laboratorio di Fisiologia e Biotecnologia dei Microrganismi, Università di Siena, I-53100 Siena, Italy

Giraud-Morin, C.; Laboratoire de Bactériologie et Epidémiologie Moléculaire, Université de Nice, Centre Hospitalier Universitaire de Nice, F-06202 Nice, France

Rossolini, G. M.; Dipartimento di Biologia Molecolare, Laboratorio di Fisiologia e Biotecnologia dei Microrganismi, Università di Siena, I-53100 Siena, Italy

Docquier, Jean-Denis ; Université de Liège - ULiège > Département des sciences de la vie > Centre d'ingénierie des protéines

Fosse, T.; Laboratoire de Bactériologie et Epidémiologie Moléculaire, Université de Nice, Centre Hospitalier Universitaire de Nice, F-06202 Nice, France

Language :

English

Title :

Genetic and biochemical characterization of TRU-1, the endogenous class C β-lactamase from Aeromonas enteropelogenes

Publication date :

2010

Journal title :

Antimicrobial Agents and Chemotherapy

ISSN :

0066-4804

eISSN :

1098-6596

Publisher :

American Society for Microbiology, Washington, United States - District of Columbia

Volume :

Issue :

Pages :

1547-1554

Peer reviewed :

Peer Reviewed verified by ORBi

Available on ORBi :

since 20 November 2020

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