Detection and purification of a catalase-peroxidase from Mycobacterium sp. Pyr-1

Abstract

A catalase-peroxidase from Mycobacterium sp. Pyr-1, a strain capable of growth on pyrene, was purified to homogeneity by anion exchange and hydroxyapatite column chromatography. The enzyme, like the M. tuberculosis T-catalase, reduced nitroblue tetrazolium in the presence of isoniazid (INH) and H₂O₂. It also oxidized 3,3′,5,5′-tetramethylbenzidine and other substrates of the catalase-peroxidase of M. tuberculosis in the presence of either tert-butyl hydroperoxide or H₂O₂. It had a UV/visible absorption spectrum (Soret peak at 406 nm) similar to that of the catalase-peroxidase of M. tuberculosis (Soret peak at 408 nm) and identical to that of the catalase-peroxidase of M. smegmatis. After electrophoresis on non-denaturing gels the enzyme showed one single protein band with both catalase and peroxidase activity, which were lost after electrophoresis on SDS-PAGE. The enzyme was inhibited by sodium azide, glutathione, 2-mercaptoethanol, and isoniazid, but not by isonicotinic acid. The optimum enzyme activity was obtained at pH 4.5 and at 25°C.

Mycobacterium, Catalase, Peroxidase, Isoniazid