Enzymatic sulfation of galactosyl- and lactosylceramides in cell lines derived from renal tubules

Abstract

1. 1.The renal cell lines, JTC-12 and MDCK, not only synthesize galactosylceramide 3-sulfate and lactosylceramide 3'-sulfate in vivo, but also contain enzymes that catalyze the transfer of sulfate to galactosylceramide and lactosylceramide in vitro. 2. 2. Concentration of cations necessary for maximum sulfotransferase activity occurred at 40 mM Ca 2+ with galactosylceramide and 15 mM Ca 2+ with lactosylceramide as the substrate. Na 2+ was also found to stimulate the sulfation of galactosylceramide, but was slightly inhibitory for the sulfation of lactosylceramide. 3. 3. The products 6f the in vitro assay mixture were characterized as galactosylceramide 3-sulfate and lactosylceramide 3'-sulfate by a variety of TLC separations. 4. 4. The apparent K m of JTC-12 cells for galactosylceramide was 17 μM, while that for lactosylceramide was 82 μM. The K m values of MDCK cells were comparable to those of JTC-12 cells. Competition studies suggested that galactosylceramide and lactosylceramide were sulfated by a single enzyme in both cell lines.