title: Properties and distribution of a metallo-β-lactamase (ALI-1) from the fish pathogen Aliivibrio salmonicida LFI1238.
abstract: OBJECTIVE
  To characterize the chromosome-encoded metallo-β-lactamase (MBL) from the psychrophilic, marine fish-pathogenic bacterium Aliivibrio salmonicida LFI1238 and check for the presence of the gene in other Aliivibrio isolates both connected to the fish-farming industry and from the environment.
  
  
  METHODS
  The MBL gene was cloned and intracellularly expressed in Escherichia coli. Kinetic parameters, NaCl dependence, pH optimum and temperature optimum were determined using purified enzyme. The VIM-2 enzyme from a Pseudomonas aeruginosa hospital isolate was used as a counterpart in comparative analysis. PCRs with degenerate MBL primers were used to screen different A. salmonicida isolates for the presence of the gene.
  
  
  RESULTS
  A. salmonicida MBL (ALI-1) is an Ambler class B β-lactamase sharing 39% and 29% amino acid identity with IMP-1 and VIM-2, respectively. ALI-1 hydrolysed all β-lactam antibiotics tested, except for the monobactam aztreonam and the penicillin piperacillin. A profound increase in activity was observed when adding NaCl to the assay mixture (60% active without addition of NaCl, increasing to 100% at 0.5 M NaCl). The increase was less noticeable for VIM-2 (100% active at 0.2 M NaCl). ALI-1 appears to be ubiquitous in nature as it is found in Aliivibrio isolates not affected by human activity.
  
  
  CONCLUSIONS
  This work provides more data for the ever-expanding MBL group of enzymes. These periplasmic enzymes are activated by addition of NaCl, and the marine enzyme is highly salt tolerant and cold active. The observed enzyme properties very likely reflect the conditions that the enzymes face in situ.
authors.affiliation: The Norwegian Structure Biology Centre (NorStruct), Department of Chemistry, University of Tromsø, N-9037 Tromsø, Norway.
authors.forename: Anders
authors.initials: A
authors.lastname: Kristiansen
authors.affiliation: The Norwegian Structure Biology Centre (NorStruct), Department of Chemistry, University of Tromsø, N-9037 Tromsø, Norway.
authors.forename: Miriam
authors.initials: M
authors.lastname: Grgic
authors.affiliation: The Norwegian Structure Biology Centre (NorStruct), Department of Chemistry, University of Tromsø, N-9037 Tromsø, Norway.
authors.forename: Bjørn
authors.initials: B
authors.lastname: Altermark
authors.affiliation: The Norwegian Structure Biology Centre (NorStruct), Department of Chemistry, University of Tromsø, N-9037 Tromsø, Norway ingar.leiros@uit.no.
authors.forename: Ingar
authors.initials: I
authors.lastname: Leiros
mesh_terms: D044165:Aliivibrio salmonicida / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D020536:Enzyme Activators / Q000378:metabolism; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D005399:Fishes; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012965:Sodium Chloride / Q000378:metabolism; D001618:beta-Lactamases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D004269:DNA, Bacterial; D020536:Enzyme Activators; D011994:Recombinant Proteins; D012965:Sodium Chloride; D001618:beta-Lactamases
keywords: A. salmonicida; MBLs; TCEP inactivation; Zn; carbapenems; psychrophilic; salt