Molecular and biochemical characterization of the natural chromosome-encoded class A beta-lactamase from Pseudomonas luteola.
Pseudomonas luteola (formerly classified as CDC group Ve-1 and named Chryseomonas luteola) is an unusual pathogen implicated in rare but serious infections in humans. A novel beta-lactamase gene, bla(LUT-1), was cloned from the whole-cell DNA of the P. luteola clinical isolate LAM, which had a weak narrow-spectrum beta-lactam-resistant phenotype, and expressed in Escherichia coli. This gene encoded LUT-1, a 296-amino-acid Ambler class A beta-lactamase with a pI of 6 and a theoretical molecular mass of 28.9 kDa. The catalytic efficiency of this enzyme was higher for cephalothin, cefuroxime, and cefotaxime than for penicillins. It was found to be 49% to 59% identical to other Ambler class A beta-lactamases from Burkholderia sp. (PenA to PenL), Ralstonia eutropha (REUT), Citrobacter sedlakii (SED-1), Serratia fonticola (FONA and SFC-1), Klebsiella sp. (KPC and OXY), and CTX-M extended-spectrum beta-lactamases. No gene homologous to the regulatory ampR genes of class A beta-lactamases was found in the vicinity of the bla(LUT-1) gene. The entire bla(LUT-1) coding region was amplified by PCR and sequenced in five other genetically unrelated P. luteola strains (including the P. luteola type strain). A new variant of bla(LUT-1) was found for each strain. These genes (named bla(LUT-2) to bla(LUT-6)) had nucleotide sequences 98.1 to 99.5% identical to that of bla(LUT-1) and differing from this gene by two to four nonsynonymous single nucleotide polymorphisms. The bla(LUT) gene was located on a 700- to 800-kb chromosomal I-CeuI fragment, the precise size of this fragment depending on the P. luteola strain.
Doublet
Benoît
B
Laboratoire des Bactéries Pathogènes Entériques, Institut Pasteur, 28 rue du Docteur Roux, 75724 Paris cedex 15, France.
Robin
Frédéric
F
Casin
Isabelle
I
Fabre
Laëtitia
L
Le Fleche
Anne
A
Bonnet
Richard
R
Weill
François-Xavier
FX
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology; D002408:Catheters, Indwelling; D002876:Chromosomes, Bacterial / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D016521:Electrophoresis, Gel, Pulsed-Field; D005798:Genes, Bacterial / Q000235:genetics*; D014644:Genetic Variation; D007525:Isoelectric Focusing; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D010802:Phylogeny; D010957:Plasmids / Q000235:genetics; D011549:Pseudomonas / Q000235:genetics*; D011552:Pseudomonas Infections / Q000382:microbiology; D011994:Recombinant Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013379:Substrate Specificity; D001618:beta-Lactamases / Q000096:biosynthesis / Q000235:genetics*
D000900:Anti-Bacterial Agents; D011994:Recombinant Proteins; D001618:beta-Lactamases