Cloning, expression, and characterization of the katG gene, encoding catalase-peroxidase, from the polycyclic aromatic hydrocarbon-degrading bacterium Mycobacterium sp strain PYR-1

A 81-kDa protein from Mycobacterium sp. strain PYR-1 was expressed in respo nse to exposure of the strain to the polycyclic aromatic hydrocarbon pyrene and recovered by two-dimensional gel electrophoresis. The N-terminal seque nce of the protein indicated that it was similar to catalase-peroxidase. An oligonucleotide probe designed from this sequence was used to screen a gen omic library of Mycobacterium sp, strain PYR-1, and a positive clone, conta ining a part of the gene encoding the 81-kDa protein, was isolated. A gene- walking technique was used to sequence the entire gene, which was identifie d as katG for catalase-peroxidase. The deduced KatG protein sequence showed significant homology to KatGII of Mycobacterium fortuitum and clustered wi th catalase-peroxidase proteins from other Mycobacterium species in a phylo genetic tree. The katG gene was expressed in Escherichia coil to produce a protein with catalase-peroxidase activity. Since the induction of this cata lase-peroxidase occurred in pyrene-induced cultures and the exposure of the se cultures to hydrogen peroxide reduced pyrene metabolism, our data sugges t that this enzyme plays a role in polycyclic aromatic hydrocarbon metaboli sm by strain PYR-1.