The Legionella (Fluoribacter) gormanii metallo-beta-lactamase: a new member of the highly divergent lineage of molecular-subclass B3 beta-lactamases.
A metallo-beta-lactamase determinant was cloned from a genomic library of Legionella (Fluoribacter) gormanii ATCC 33297(T) constructed in the plasmid vector pACYC184 and transformed into Escherichia coli DH5alpha, by screening for clones showing a reduced susceptibility to imipenem. The product of the cloned determinant, named FEZ-1, contains a 30-kDa polypeptide and exhibits an isoelectric pH of 7.6. Sequencing revealed that FEZ-1 is a molecular-class B beta-lactamase which shares the closest structural similarity (29.7% of identical residues) with the L1 enzyme of Stenotrophomonas maltophilia, being a new member of the highly divergent subclass B3 lineage. All the residues that in L1 are known to be directly or indirectly involved in coordination of the zinc ions were found to be conserved also in FEZ-1, suggesting that the geometry of zinc coordination in the active site of the latter enzyme is identical to that of L1. Unlike L1, however, FEZ-1 appeared to be monomeric in gel permeation chromatography experiments and exhibited a distinctive substrate specificity with a marked preference for cephalosporins and meropenem. The properties of FEZ-1 overall resembled those of a beta-lactamase previously purified from the same strain of L. gormanii (T. Fujii, K. Sato, K. Miyata, M. Inoue, and S. Mitsuhashi, Antimicrob. Agents Chemother. 29:925-926, 1986) and are as yet unique among class B enzymes, reinforcing the notion that considerable functional heterogeneity can be encountered among members of this class. A system for overexpression of the bla(FEZ-1) gene in E. coli, based on the T7 phage promoter, was also developed.
Boschi
L
L
Dipartimento di Biologia Molecolare, Sezione di Microbiologia, Università di Siena, I-53100 Siena, Italy.
Mercuri
P S
PS
Riccio
M L
ML
Amicosante
G
G
Galleni
M
M
Frère
J M
JM
Rossolini
G M
GM
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D005798:Genes, Bacterial; D007875:Legionella / Q000201:enzymology / Q000235:genetics*; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D001618:beta-Lactamases / Q000235:genetics* / Q000378:metabolism
D001618:beta-Lactamases