Simultaneous azo-coupling method for an estrogen sulfatase in human tissues.
A simultaneous azo-coupling method for the histochemical localization of d-equilenin sulfatase is described. d-Equilenin is a natural estrogenic steroid hormone, and its sulfuric acid ester was synthesized. It was found that the d-equilenin liberated during hydrolysis of d-equilenin sulfate by tissue sulfatase could be coupled with a diazonium salt to produce a purple precipitate indicating enzyme activity. d-Equilenin sulfatase was found in human tissues, but not in tissues of the rat. The optimum substrate concentration was 0.8 mM, activity was demonstrable over the wide pH range 5.0-8.0. Enzyme activity localized diffusely in the cytoplasm in optimally fixed specimens. Enzyme activity was also fairly well demonstrable in unfixed cryostat sections. Enzyme activity was completely inhibited by 0.1 M phosphate, 1 mM sodium tetraborate, 1 mM p-nitrophenyl sulfate and by 2 mM p-nitrocatechol sulfate. Estrone sulfate at concentration 0.8 mM had no effect, but at 4 mM caused marked inhibition of the reaction. At the same concentrations dehydroepiandrosterone sulfate did not inhibit the reaction. The chemical properties and tissue localizations of d-equilenin sulfatase differed from the properties of arylsulfatases A, B and C and other steroid sulfatases reported previously in the literature.
D015068:17-Ketosteroids / Q000032:analysis*; D000313:Adrenal Medulla / Q000201:enzymology; D001940:Breast / Q000201:enzymology; D003979:Diazonium Compounds; D004717:Endometrium / Q000201:enzymology; D004855:Equilenin / Q000032:analysis*; D005260:Female; D006651:Histocytochemistry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007684:Kidney Tubules / Q000201:enzymology; D008297:Male; D010053:Ovary / Q000201:enzymology; D011247:Pregnancy; D012394:Rosaniline Dyes; D012867:Skin / Q000201:enzymology; D013429:Sulfatases / Q000378:metabolism*; D013997:Time Factors; D014052:Toluidines; D014327:Trophoblasts / Q000201:enzymology
D015068:17-Ketosteroids; D003979:Diazonium Compounds; D012394:Rosaniline Dyes; D014052:Toluidines; C020747:Fast Blue B; C005409:pararosaniline; D013429:Sulfatases; C020121:estrone sulfatase; D004855:Equilenin