ISOLATION AND PARTIAL-PURIFICATION OF MITOCHONDRIAL AND CYTOSOLIC RHODANESE FROM LIVER OF NORMAL AND P-DIMETHYLAMINOAZOBENZENE TREATED MICE

Rhodanese (thiosulfate:cyanide sulfurtransferase, E.C, 2.8.1.1), an en zyme involved in heme regulation, showed distinctive mitochondrial and cytoplasmic activities in several models of tumorigenesis. To investi gate the basis for these differences, the enzyme was partly purified a nd characterized from the mitochondrial and cytosolic liver fraction o f mice treated with the carcinogen p-dimethyl-aminoazobenzene (DAB). A Linear relationship between incubation time and specific activity was observed up to about 30 min for cytosolic enzyme and 15 min for mitoc hondrial enzyme irrespective of whether or not the enzyme was derived from treated or untreated animals, The optimum incubation temperature was 3 degrees C for the enzyme of both fractions in control animals an d 30 degrees C for treated animals in both cases, In control and DAB t reated animals the cytoplasmic rhodanese exhibited a maximum at a lowe r pH than for the mitochondrial enzyme, The enzyme showed typical Mich aelis-Menten behavior with cyanide inhibition at concentrations higher than 25 mM for controls and 10 mM for treated animals for both fracti ons and thiosulfate inhibition at concentrations higher than 100 mM in all cases studied, K-m values of 190 and 65.66 mM were obtained for t hiosulfate and 6.37 and 9.79 mM for cyanide for both mitochondrial and cytosolic fractions of control animals; while K-m values of 31.75 and 4.58 mM were obtained for thiosulfate and 0.61 and 1.11 mM for cyanid e in both fractions of treated animals. We demonstrated differences in the kinetics for rhodanese derived from mitochondrial and cytoplasmic fractions of livers taken from tumor bearing mice, These differences might provide an explanation for the abnormalities of heme synthesis p reviously reported during hepatocarcinogenesis.