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Acceptor Substrate Specificity of a Cloned G D3 Synthase that Catalyzes the Biosynthesis of Both G D3 and G D1c /G T1a /G Q1b

1996; Wiley; Volume: 238; Issue: 3 Linguagem: Inglês

10.1111/j.1432-1033.1996.0647w.x

1432-1033

Kiyomitsu Nara, Yumiko Watanabe, Ikuo Kawashima, Tadashi Tai, Yoshitaka Nagai, Yutaka Sanai,

Enzyme Structure and Function

To address the role of α2,8‐sialyltransferase (G D3 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 G D3 from G M3 , and sialyltransferase V (SAT V) catalyzes the production of G D1c /G T1a /G Q1b from G M1b /G D1a /G T1b . However, acceptor specificity of the cloned G D3 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(α2–3)Gal structure of the carbohydrate moiety, which includes G M3 , G M1b , G D1a , and G T1b as exogenous substrates. Kinetic data also showed that the enzyme was able to utilize both G M3 and G M1b /G D1a /G T1b as acceptor substrates. These data indicate that the enzyme catalyzes the formation of not only G D3 but also G D1c , G T1a , and G Q1b in vitro. Furthermore, by transfection of the cloned human α2,8‐sialyltransferase cDNA, transient and stable expression of G T1 a and G Q1b was 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.