title: CAU-1, a subclass B3 metallo-beta-lactamase of low substrate affinity encoded by an ortholog present in the Caulobacter crescentus chromosome.
abstract: The sequenced chromosome of Caulobacter crescentus CB15 encodes a hypothetical protein that exhibits significant similarity (30 to 35% identical residues) to metallo-beta-lactamases of subclass B3. An allelic variant of this gene (divergent by 3% of its nucleotides) was cloned in Escherichia coli from C. crescentus type strain DSM4727. Expression studies confirmed the metallo-beta-lactamase activity of its product, CAU-1. The enzyme produced in E. coli was purified by two ion-exchange chromatography steps. CAU-1 contains a 29-kDa polypeptide with an alkaline isoelectric pH (> 9), and unlike the L1 enzyme of Stenotrophomonas maltophilia, the native form is monomeric. Kinetic analysis revealed a preferential activity toward penicillins, carbapenems, and narrow-spectrum cephalosporins, while oxyimino cephalosporins were poorly or not hydrolyzed. Affinities for the various beta-lactams were poor overall (K(m) values were always > 100 microM and often > 400 microM). The interaction with divalent ion chelators appeared to occur by a mechanism similar to that prevailing in other members of subclass B3. In C. crescentus, the CAU-1 enzyme is produced independently of beta-lactam exposure and, interestingly, the bla(CAU) determinant is bracketed by three other genes, including two genes encoding enzymes involved in methionine biosynthesis and a gene encoding a putative transcriptional regulator, in an operon-like structure. The CAU-1 enzyme is the first example of a metallo-beta-lactamase in a member of the alpha subdivision of the class Proteobacteria:
authors.affiliation: Dipartimento di Biologia Molecolare, Sezione di Microbiologia, Università di Siena, 53100 Siena, Italy.
authors.forename: Jean-Denis
authors.initials: JD
authors.lastname: Docquier
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Pantanella
authors.forename: Francesco
authors.initials: F
authors.lastname: Giuliani
authors.forename: Maria Cristina
authors.initials: MC
authors.lastname: Thaller
authors.forename: Gianfranco
authors.initials: G
authors.lastname: Amicosante
authors.forename: Moreno
authors.initials: M
authors.lastname: Galleni
authors.forename: Jean-Marie
authors.initials: JM
authors.lastname: Frère
authors.forename: Karen
authors.initials: K
authors.lastname: Bush
authors.forename: Gian Maria
authors.initials: GM
authors.lastname: Rossolini
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D016934:Caulobacter / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics*; D002614:Chelating Agents / Q000494:pharmacology; D002850:Chromatography, Gel; D002876:Chromosomes, Bacterial / Q000235:genetics*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000096:biosynthesis / Q000235:genetics; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D021241:Spectrometry, Mass, Electrospray Ionization; D047090:beta-Lactams
chemical_list: D000900:Anti-Bacterial Agents; D002614:Chelating Agents; D004269:DNA, Bacterial; D011994:Recombinant Proteins; D047090:beta-Lactams