Isolation and partial purification of mitochondrial and cytosolic rhodanese from liver of normal and p-dimethylaminoazobenzene treated mice

ELBA VÁZQUEZ; CÉSAR POLO; GUSTAVO STEDILE; CAROLINA SCHEBOR; EDUARDO KARAHANIAN; ALCIRA BATLLE

INTERNATIONAL JOURNAL OF BIOCHEMISTRY AND CELLULAR BIOLOGY

Elsevier

Año: 1995 vol. 27 p. 523 - 529

1357-2725

Rhodanese (thiosulfste:cyanide sulfurtransferase, E.C. 2.8.1.1), an enzyme involved in hemeregulatbn, showed disthctive mitocbondrial and cytoplasmic activities in several models Oftumorigenesis. To investigate the basis for thw dilferences, the enzyme was partly puri8ed andcharacteri& from the mitochondriai and cytosolic liver fraction of mice treated with thecarchvogeu p-dimethyl-amhmasobenzene (DAB).A liuear r&tiuss&lp between hrcubation time and specitk activity was observed up to about30 min for cytosolic enuyme and 15 min for mitodmudrial enzyme irrespective of whet&r or notthe enzyme was derived from treated or untreated animals. The optimum hmubation temperaturewas 3OC for the ensyme of botb fractious in control a&u& and 38°C for treated animals in bothcases. In control and DAB treated animals the cytophrsmic rhodanese exhibited a maximum ata lower pH tbau for the mitocbondrial enzyme.The enzyme showed typical Michaelis-Menten behavior with cyanide inhibition at coucentrationshigher than 25 mM for controls and 10 mM for treated animals for both fractions andthiosulfate hrhi~tiou at concentrations higher than 100 mM in all cases studied. Km values of 190and 65.66mM were obtained for thlosulfate and 6.37 and 9.79mM for cyanide for bothmitochombial and cytosollc fractions of control anhnalq while Km values of 31.75 and 4.58 mMwere obtained for thiosulfate and 0.61 and 1.11 mM for cyanide in both fractions of treatedanimals.We demonstrated dllferences in the kinetics for rhodanese derived from mito&oudrial andcytoplPsmic fractions of livers taken from tumor bearing mice. These diierences might providean explanation for the abnormalities of heme synthesis previously reported during bepatocarcinogenesis.