Acceptor substrate specificity of a cloned GD3 synthase that catalyzes the biosynthesis of both GD3 and GD1c/GT1a/GQ1b.
To address the role of alpha2,8-sialyltransferase (GD3 synthase) in the biosynthesis of gangliosides, we examined the substrate specificity of the enzyme. In the ganglioside synthesis pathway, it has been generally accepted that sialyltransferase II (SAT II) catalyzes the production of GD3 from GM3, and sialyltransferase V (SAT V) catalyzes the production of GD1c/GT1a/GQ1b from GM1h/GD1a/GT1b. However, acceptor specificity of the clones GD3 synthase that was isolated from human melanoma cells [Nara, K., Watanabe, Y., Maruyama, K., Kasahara, K., Nagai. Y. & Sanai, Y. (1994) Proc. Natl Acad. Sci. USA 91, 7952-7956] has revealed that this enzyme utilized the gangliosides containing the terminal Sia(alpha2-3)Gas structure of the carbohydrate moiety, which includes GM3, GM1b, GD1a and GT1B as exogenous substrates. Kinetic data also showed that the enzyme was able to utilize both GM3 and GM1b/GD1a/GT1b as acceptor substrates. These data indicate that the enzyme catalyzes the formation of not only GD3 but also GD1c, GT1a, and GQ1B in vitro. Furthermore, by transfection of the cloned human alpha2,8-sialyltransferase cDNA, transient and stable expression of GT1a and GQ1b wa also observed in COS-7 cells and Swiss 3T3 cells that originally lacked SAT II and SAT V activities. These observations indicate that the enzyme has both SAT II and SAT V activities in vivo.
Nara
K
K
Department of Biochemical Cell Research, Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan.
Watanabe
Y
Y
Kawashima
I
I
Tai
T
T
Nagai
Y
Y
Sanai
Y
Y
D016475:3T3 Cells; D000818:Animals; D002240:Carbohydrate Sequence; D005732:Gangliosides / Q000096:biosynthesis* / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012799:Sialyltransferases / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D014162:Transfection
D005732:Gangliosides; D011994:Recombinant Proteins; D012799:Sialyltransferases; C043662:alpha-N-acetylneuraminate alpha-2,8-sialyltransferase