How does hexachlorobenzene treatment affect liver uroporphyrinogen decarboxylase?
The aims of the present work were: (1) to investigate whether the strong decrease of liver uroporphyrinogen decarboxylase (UroD) activity observed in experimental porphyria cutanea tarda is due to alteration of the enzymatic protein and (2) to improve the knowledge about the normal liver enzyme. With these purposes, several physicochemical studies for enzymatic characterization were carried out comparatively on the 12-fold purified liver enzyme of both normal and hexachlorobenzene porphyric rat. The study shows that the enzyme from porphyric rats has a higher activation energy, lower reactivity index and lower optimum pH than the normal one. In addition, it did not reach the Vmax at any of the substrate concentrations assayed (up to 28 microM uroporphyrinogen III), while the normal enzyme reached the plateau around 14 microM. The porphyric enzyme appears to be more protected than the normal against the inhibitory action of several metals, particularly Cu2+ and Pb2+, and against thermal inactivation. Zn2+ did not affect enzymatic activity, whereas Cu2+, Hg2+, Fe2+, Pb2+, and Cd2+ lowered the activities of both normal and porphyric enzyme in a dose-related way. It was also observed that the larger the atomic radius in its hydrated state, the lower the effect of the metal. Neither glutathione nor dithiothreitol significantly altered enzymatic activity in the range of concentrations assayed. beta-Mercaptoethanol had diverse effects, as regards both the concentration assayed and the enzymatic sample used. Assays with cystine showed a dual behaviour of both normal and porphyric enzymatic activity. Western blots for both preparations revealed a single band (65 kDa) with a similar intensity. This study show that hexachlorobenzene treatment modifies the physicochemical properties of liver UroD leading to a sharp decrease of its activity, without affecting its antigenic reactivity probably as a consequence of changes at the conformational level promoted by the binding of its reported inhibitor.
Chaufan
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Laboratorio de Porfirias Experimentales y Metabolismo del Hemo, Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina.
Ríos de Molina
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MC
San Martín de Viale
L C
LC
D000818:Animals; D000941:Antigens / Q000276:immunology; D005260:Female; D005659:Fungicides, Industrial / Q000378:metabolism* / Q000494:pharmacology; D006581:Hexachlorobenzene / Q000378:metabolism* / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000187:drug effects / Q000201:enzymology*; D051381:Rats; D017208:Rats, Wistar; D013696:Temperature; D014575:Uroporphyrinogen Decarboxylase / Q000378:metabolism*
D000941:Antigens; D005659:Fungicides, Industrial; D006581:Hexachlorobenzene; D014575:Uroporphyrinogen Decarboxylase