Steroid sulfatase from the microsomal fraction of rat liver exhibits properties which are suggestive of an allosteric type of enzyme. The kinetics of dehydroisoandrosterone sulfate cleavage are normal while the cleavage of cholesterol sulfate presents a cooperative effect beyond 0.8 × 10−6 Mat the protein concentration used. This abnormal kinetic behavior is normalized by the addition of an analogue of this substrate, sodium lauryl sulfate. This analogue elicits a biphasic effect on the cleavage of cholesterol sulfate while the cleavage of dehydroisoandrosterone sulfate and pregnenolone sulfate is inhibited at all of the concentrations tested.Using polyacrylamide gel electrophoresis in the presence of sodium lauryl sulfate, the molecular weight of what is believed to be the monomeric form of the enzyme was estimated to be approximately 23 000.Steriod sulfatase fromHelix pomatiacleaves the sulfate of dehydroisoandrosterone while cholesterol sulfate remains intact. This enzyme was found to have a molecular weight near or below 50 000 and was isolated as a single band of sulfatase activity by polyacrylamide gel electrophoresis.