Cloning, sequence analysis; and expression of the Pseudomonas putida 33/1 1H-3-hydroxy-4-oxoquinoline 2,4-dioxygenase gene, encoding a carbon monoxide forming dioxygenase

1H-3-hydroxy-4-oxoquinoline 2,4-dioxygenase (Qdo) from the 1H-4-oxoquinolin e utilizing Pseudomonas putida strain 33/1, which catalyzes the cleavage of 1H-3-hydroxy-4-oxoquinoline to carbon monoxide and N-formylanthranilate, i s devoid of any transition metal ion or other cofactor and thus represents a novel type of ring-cleavage dioxygenase. Gene qdo was cloned and sequence d. Its overexpression in Escherichia coli yielded recombinant His-tagged Qd o which was catalytically active. Qdo exhibited 36% and 16% amino acid iden tity to 1H-3-hydroxy-4-oxoquinaldine 2,4-dioxygenase (Hod) and atropinester ase (a serine hydrolase), respectively. Qdo as well as Hod possesses a SXSH G motif, resembling the motif GXSXG of the serine hydrolases which comprise s the active-site nucleophile (X=arbitrary residue). (C) 1999 Elsevier Scie nce B.V. All rights reserved.