MOLECULAR-CLONING, EXPRESSION, AND ENZYMATIC CHARACTERIZATION OF RABBIT HYDROXYSTEROID SULFOTRANSFERASE AST-RB2 (ST2A8)

Cytosolic sulfotransferases, which consist of at least three gene fami lies, play a major role in activation and detoxification of both endog enous and exogenous chemicals. We recently purified a rabbit sulfotran sferase, AST-RB2, showing high activities to both hydroxysteroids and amines. To characterize this enzyme, a rabbit cDNA library was screene d using anti-AST-RB2 antibodies, The isolated cDNA was judged to encod e AST-RB2 (ST2A8) based on the amino acid sequences of peptide fragmen ts obtained from purified AST-RB2. The cDNA showed high similarity to other mammalian hydroxysteroid sulfotransferases (ST2) at the amino ac id level (58-68%), but low similarity to aryl sulfotransferases (ST1) (less than 37%). The protein expressed in Escherichia coli catalyzed s ulfation of typical ST2 substrates, Therefore, ST2A8 was judged to bel ong to the ST2 family from both its primary structure and substrate sp ecificity. The ST2A8 protein expressed in E. coli clearly differed fro m rat ST2A1 and ST2A2 on its localization (cytosol/insoluble fraction ratio), ST2A8 had no activity to lithocholate, but showed the highest catalysis on dehydroepiandrosterone and testosterone among the four fo rms (ST2A1, ST2A2, ST2A3, and ST2A8), indicating a clear difference be tween ST2A forms in substrate specificity to endogenous chemicals.