The Aeromonas hydrophila cphA gene: molecular heterogeneity among class B metallo-beta-lactamases.
An Aeromonas hydrophila gene, named cphA, coding for a carbapenem-hydrolyzing metallo-beta-lactamase, was cloned in Escherichia coli by screening an Aeromonas genomic library for clones able to grow on imipenem-containing medium. From sequencing data, the cloned cphA gene appeared able to code for a polypeptide of 254 amino acids whose sequence includes a potential N-terminal leader sequence for targeting the protein to the periplasmic space. These data were in agreement with the molecular mass of the original Aeromonas enzyme and of the recombinant enzyme produced in E. coli, evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of crude beta-lactamase preparations followed by renaturation treatment for proteins separated in the gel and localization of protein bands showing carbapenem-hydrolyzing beta-lactamase activity by a modified iodometric technique. The deduced amino acid sequence of the CphA enzyme showed regions of partial homology with both the beta-lactamase II of Bacillus cereus and the CfiA beta-lactamase of Bacteroides fragilis. Sequence homologies were more pronounced in the regions encompassing the amino acid residues known in the enzyme of B. cereus to function as ligand-binding residues for the metal cofactor. The CphA enzyme, however, appeared to share a lower degree of similarity with the two other enzymes, which, in turn, seemed more closely related to each other. These results, therefore, suggest the existence of at least two molecular subclasses within molecular class B metallo-beta-lactamases.
Massidda
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Dipartimento di Biologia Molecolare, Sezione Microbiologia, Universitá degli Studi di Siena, Italy.
Rossolini
G M
GM
Satta
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G
D000333:Aeromonas / Q000201:enzymology / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D015780:Carbapenems / Q000494:pharmacology*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005786:Gene Expression Regulation; D005798:Genes, Bacterial; D014644:Genetic Variation; D006868:Hydrolysis; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D001618:beta-Lactamases / Q000235:genetics* / Q000502:physiology
D015780:Carbapenems; D004269:DNA, Bacterial; D001618:beta-Lactamases