New assay for steroid sulfatase (EC 3.1.6.2) and its application for studies of human placental and skin sulfatase.
A new, simple, fast and highly practicable sulfatase assay and its application is described. Sterol sulfatase sulfohydrolase (EC 3.1.6.2) activity is determined by a two-phase scintillation technique separating the unreacted [4-14C]dehydroepiandrosterone sulfate from carbon-14-labeled products. The principle of the separation relies on the limited emulsifying capacity of the dioxane-based scintillation solution for water and the different partition of dehydroepiandrosterone sulfate and sulfate-free steroid products between the scintillation fluid and the aqueous phase as recently applied for determination of aromatase activity [1]. [7-3H]Dehydroepiandrosterone sulfate can also be used as a substrate for this assay. This test was applied to studies of microsomal sulfatase prepared from human term placenta and to the detection of sulfatase activity in human skin biopsies. Using placental microsomes, the Km of dehydroepiandrosterone sulfate was determined to be 5.0 X 10(7)M. Sulfatase activity in frozen scrotal skin was found to be 2-3 fold than with vaginal skin. Using an incubation time of 24h/skin sulfatase can be detected in biopsies as small as 2.5 mm2. The sulfatase assay can be applied for routine detection of human placental sulfatase deficiency and, furthermore, the application of this assay has to be demonstrated for the analysis of sulfatase activity in patients with congenital ichthyosis (X-chromosomal, recessive type).
D002250:Carbon Radioisotopes; D005260:Female; D006801:Humans; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology*; D010920:Placenta / Q000201:enzymology*; D011247:Pregnancy; D011865:Radioisotope Dilution Technique; D012867:Skin / Q000201:enzymology*; D043266:Steryl-Sulfatase; D013429:Sulfatases / Q000032:analysis* / Q000378:metabolism
D002250:Carbon Radioisotopes; D013429:Sulfatases; D043266:Steryl-Sulfatase