Purification of chondroitin 6-sulfotransferase secreted from cultured chick embryo chondrocytes.
Chondroitin 6-sulfotransferase, which transfers sulfate from 3'-phosphoadenylyl sulfate to position 6 of N-acetylgalactosamine in chondroitin, was purified 1,430-fold to apparent homogeneity with a 22% yield from the serum-free culture medium of chick embryo chondrocytes by affinity chromatography on heparin-Sepharose CL-6B, wheat germ agglutinin-agarose, and 3',5'-ADP-agarose. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified enzyme showed a single broad protein band with an apparent molecular weight of 75,000. Since the purified enzyme has an apparent molecular weight of 160,000 as judged by gel chromatography on Superose 12, the active form of chondroitin 6-sulfotransferase may be a dimer. The purified enzyme transferred sulfate to chondroitin, chondroitin sulfate, and corneal keratan sulfate. Chondroitin sulfate E from squid cartilage, dermatan, sulfate, and heparan sulfate hardly served as acceptors of the sulfotransferase. The sulfated product derived from keratan sulfate was degraded by keratanase but not by chondroitinase ABC.
Habuchi
O
O
Department of Life Science, Aichi University of Education, Kariya, Japan.
Matsui
Y
Y
Kotoya
Y
Y
Aoyama
Y
Y
Yasuda
Y
Y
Noda
M
M
D000818:Animals; D002356:Cartilage / Q000201:enzymology*; D002478:Cells, Cultured; D002642:Chick Embryo; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D016895:Culture Media, Serum-Free; D004591:Electrophoresis, Polyacrylamide Gel; D013379:Substrate Specificity; D015238:Sulfotransferases / Q000302:isolation & purification* / Q000378:metabolism; D000097811:Carbohydrate Sulfotransferases
D016895:Culture Media, Serum-Free; D015238:Sulfotransferases; D000097811:Carbohydrate Sulfotransferases; C019893:chondroitin 4-sulfotransferase