MOLECULAR-CLONING AND EXPRESSION OF CDNA-ENCODING HUMAN 3'-PHOSPHOADENYLYLSULFATE-GALACTOSYLCERAMIDE 3'-SULFOTRANSFERASE

We have isolated a cDNA clone encoding human 3'-phosphoadenylylsulfate :galactosylceramide 3'-sulfotransferase (EC 2.8.2.11). Degenerate olig onucleotides, based on amino acid sequence data for the purified enzym e, were used as primers to amplify fragments of the gene from human re nal cancer cell cDNA by the polymerase chain reaction method. The ampl ified cDNA fragment was then used as probe to screen a human renal can cer cell cDNA library. The isolated cDNA clone contained an open readi ng frame encoding 423 amino acids including all of the peptides that w ere sequenced. The deduced amino acid sequence predicts a type II tran smembrane topology and contains two potential N-glycosylation sites. T here is no significant homology between this sequence and either the s ulfotransferases cloned to date or other known proteins. Northern blot analysis demonstrated that a 1.9-kilobase mRNA was unique to renal ca ncer cells. When the cDNA was inserted into the expression vector pSVK 3 and transfected into COS-l cells, galactosylceramide sulfotransferas e activity in the transfected cells increased from 8- to 16-fold over that of controls, and the enzyme product, sulfatide, was expressed on the transformed cells.