Steroid sulphatase: expression, isolation and inhibition for active-site identification studies.
Steroid sulphatase, which can hydrolyse 3-hydroxysteroid sulphates, has important roles in several physiological and pathological processes. A number of steroid sulphatase inhibitors have now been developed, of which the most potent to date is oestrone-3-O-sulphamate (EMATE). This inhibitor inactivates steroid sulphatase in an irreversible, time- and concentration-dependent manner. In order to be able to use a radiolabelled derivative of EMATE to study the active site, it will be essential to prepare the steroid sulphatase in a pure form. For this, attempts have been made to express the protein, using the steroid sulphatase cDNA, in the pGEX2T expression system and also to express a mutant form of the protein, in which the putative membrane-spanning domain was deleted, in CHO cells. In addition, a soluble steroid sulphatase has been identified from the snail Helix pomatia. This steroid sulphatase is inhibited by EMATE in an irreversible manner, similar to the human steroid sulphatase and appears to possess a histidine residue at its active site. The expression and/or isolation of a steroid sulphatase, in conjunction with the use of a radiolabelled derivative of EMATE should allow important new information about the active site of this enzyme and the mechanism of its inactivation to be obtained.
Purohit
A
A
Unit of Metabolic Medicine, Imperial College School of Medicine at St. Mary's, London, UK.
Potter
B V
BV
Parker
M G
MG
Reed
M J
MJ
D000818:Animals; D001192:Arylsulfatases / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000302:isolation & purification*; D001665:Binding Sites; D016466:CHO Cells; D006224:Cricetinae; D004791:Enzyme Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D004970:Estrone / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D043266:Steryl-Sulfatase
D004791:Enzyme Inhibitors; C088768:estrone-3-O-sulfamate; D004970:Estrone; D001192:Arylsulfatases; D043266:Steryl-Sulfatase