Purification and properties of goat liver catalase: two pH optima.

ABSTRACT

Goat liver catalase (EC 1.11.1.6) has been purified to homogeneity using the techniques of ammonium sulfate fractionation, DEAE-cellulose chromatography and gel-filtration through Ultrogel AcA-34 involving two alternating steps of column chromatography. The homogeneity of the purified enzyme was tested by native and sodium dodecyl sulfate polyacrylamide gel electrophoresis, immunodiffusion and immunoelectrophoresis. The enzyme is a tetramer having a subunit molecular weight of 58,000 +/- 3000, contains six sulfhydryl groups per mole of the enzyme and shows pH optima at pH 6.8 and 7.7. The kinetic data show no cooperativity between the substrate binding sites. Tryptophan, indoleacetic acid, cysteine, formaldehyde and sodium azide inhibit the enzyme non-competitively with Ki values of 4 +/- 1, 2.5 +/- 0.8, 6 +/- 1.5, 0.48 +/- 0.15 and 0.0013 +/- 0.0003 mM, respectively. Sulfhydryl group binding agents as well as thiol reagents inhibit the enzyme activity.