CMP-NeuAc:Gal beta 1-->4GlcNAc alpha 2-->6sialyltransferase catalyzes NeuAc transfer to glycolipids.
Using mammalian gene-overexpression system, in vitro catalytic activities of CMP-NeuAc:Gal beta 1-->4GlcNAc alpha 2-->6sialytransferase on glycosphinogolipid acceptors were analyzed. We transfected the mammalian expression vector containing the cDNA that was cloned from Daudi cells into COS-1 cells, and selected monoclonal transfectants in the presence of G418. Although the transfected alpha 2-->6sialytransferase can catalyze NeuAc transfer onto glycoprotein acceptors more than glycolipids based on kinetic analyses, the substantial synthesis of IV6NeuAc-nLcOse4Cer was observed and the activities were 7- to 9-times higher in the transfected cells than in the mock transfectants. In addition, the transfected COS-1 cells with alpha 2-->6sialytransferase cDNA were revealed to contain a higher amount of ganglioside that has the terminal NeuAc alpha 2-->6Gal sequence in the in situ situation than the mock transfectants. These results using transfectants, together with those using the purified enzyme protein, suggest that the alpha 2-->6sialytransferase enzyme from Daudi cells can also catalyze NeuAc transfer in alpha 2-->6 linkage onto glycosphingolipid acceptors.
Nakamura
M
M
Division of Hemopoiesis, Jichi Medical School, Tochigi, Japan.
Tsunoda
A
A
Yanagisawa
K
K
Furukawa
Y
Y
Kikuchi
J
J
Iwase
S
S
Sakai
T
T
Larson
G
G
Saito
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D005732:Gangliosides / Q000737:chemistry / Q000378:metabolism; D006017:Glycolipids / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D016133:Polymerase Chain Reaction; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D012799:Sialyltransferases / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D014162:Transfection; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase
D017931:DNA Primers; D018076:DNA, Complementary; D005732:Gangliosides; D006017:Glycolipids; D011993:Recombinant Fusion Proteins; D012799:Sialyltransferases; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase