caa a22 4500 397558287 CHVBK 20180308164809.0 cr unu---uuuuu 161202e199603 xx s 000 0 eng 10.1093/oxfordjournals.jbchem.a021258 doi (NATIONALLICENCE)oxford-10.1093/oxfordjournals.jbchem.a021258 Purification and Characterization of 3′-Phosphoadenosine-5′-Phosphosulfate: GalCer Sulfotransferase from Human Renal Cancer Cells [Elektronische Daten] [Koichi Honke, Miwako Yamane, Atsushi Ishii, Takahiko Kobayashi, Akira Makita] We have purified 3′-phosphoadenosine-5′-phosphosulfate: GalCer sulfotransferase [EC 2.8.2.11] from a human renal cancer cell line SMKT-R3 through a combination of affinity chromatographies using galactosylsphingosine, 3′,5′-bisphosphoadenosine and heparin as ligands. The purified sulfotransferase showed a specific activity of 1.2 μmol/min/mg, which is 300 times more than the highest activity among the enzyme preparations purified so far from other sources. Homogeneity of the purified sulfotransferase was supported by the facts that the enzyme preparation showed a single protein band with an apparent molecular mass of 54 kDa on reducing SDS-PAGE and that protein bands coincided with the enzyme activity on both native PAGE and nonreducing SDS-PAGE. GalCer was the best acceptor for the purified enzyme. LacCer, GalAAG, and GalDG were also good acceptors. GlcCer, Gg3Cer, Gg4Cer, Gb4Cer, and nLc4Cer did serve as acceptors although the relative activities were low. On the other hand, the enzyme could not act on Gb3Cer, which possesses α-galactosid at the nonreducing terminus. Neither galactose nor lactose served as an acceptor. These observations suggest that the sulfotransferase prefers β-glycoside, especially β-galactoside, at the nonreducing termini of sugar chains attached to a lipid moiety. © by the Journal of Biochemistry Regular Papers nationallicence affinity chromatography nationallicence biosynthesis of glycosphingolipids nationallicence renaturation nationallicence substrate specificity nationallicence sulfatide nationallicence Honke Koichi Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut Yamane Miwako Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut Ishii Atsushi Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut Kobayashi Takahiko Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut Makita Akira Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut The Journal of Biochemistry Oxford University Press 119/3(1996-03), 421-427 0021-924X 119:3<421 1996 119 jbchem https://doi.org/10.1093/oxfordjournals.jbchem.a021258 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1093/oxfordjournals.jbchem.a021258 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Honke Koichi Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut NATIONALLICENCE 700 1- Yamane Miwako Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut NATIONALLICENCE 700 1- Ishii Atsushi Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut NATIONALLICENCE 700 1- Kobayashi Takahiko Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut NATIONALLICENCE 700 1- Makita Akira Biochemistry Laboratory, Cancer Institute, Hokkaido University School of Medicine, Kita-ku, Sapporo 060 aut NATIONALLICENCE 773 0- The Journal of Biochemistry Oxford University Press 119/3(1996-03), 421-427 0021-924X 119:3<421 1996 119 jbchem Metadata rights reserved CC BY-NC-4.0 http://creativecommons.org/licenses/by-nc/4.0 nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-oxford