Escherichia coli chromosomal mutations that permit direct cloning of the Bacteroides fragilis metallo-beta-lactamase gene, ccrA.
The class B, metallo-beta-lactamase genes ccrA (carbapenem- and cephamycin resistance) from three Bacteroides fragilis isolates--QMCN3, QMCN4, and TAL3636--were cloned and expressed in Escherichia coli. Cloning of the genes, by selecting for ampicillin resistance, was facilitated by two classes of Escherichia coli chromosomal mutations which resulted in at least a 5-10-fold increase in metallo-beta-lactamase enzymatic activity. The observed increase in enzymatic activity is due to either increased translation of the ccrA gene or an effect on localization or stability of the protein. Comparison of the DNA sequences of the three ccrA genes revealed that their protein-coding sequences shared greater than 97% DNA sequence identity. However, the 5' upstream sequence for the TAL3636 ccrA gene was unrelated to that of the other two genes.
Rasmussen
B A
BA
Molecular Biology Section, American Cyanamid Company, Pearl River, New York 10965.
Gluzman
Y
Y
Tally
F P
FP
D000595:Amino Acid Sequence; D000668:Ampicillin Resistance; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001441:Bacteroides fragilis / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D015032:Zinc / Q000378:metabolism; D001618:beta-Lactamases / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D001618:beta-Lactamases; C063912:carbapenemase; D015032:Zinc