Molecular cloning and functional expression of two members of mouse NeuAc alpha 2,3Gal beta 1,3GalNAc GalNAc alpha 2,6-sialyltransferase family, ST6GalNAc III and IV

Two cDNA clones encoding NeuAc alpha 2,3Gal beta 1,3GalNAc GalNAc alpha 2,6 -sialyltransferase have been isolated from mouse brain cDNA libraries. One of the cDNA clones is a homologue of previously reported rat ST6GalNAc III according to the amino acid sequence identity (94.4%) and the substrate spe cificity of the expressed recombinant enzyme, while the other cDNA clone in cludes an open reading frame coding for 302 amino acids. The deduced amino acid sequence is not identical to those of other cloned mouse sialyltransfe rases, although it shows the highest sequence similarity with mouse ST6GalN Ac III (43.0%). The expressed soluble recombinant enzyme exhibited activity toward NeuAc alpha 2, 3Gal beta 1,3GalNAc, fetuin, and GM1b, while no sign ificant activity was detected toward Gal beta 1,3GalNAc or asialofetuin, or the other glycoprotein substrates tested. The sialidase sensitivity of the C-14-sialylated residue of fetuin, which was sialylated by this enzyme wit h CMP-[C-14]NeuAc, was the same as that of ST6GalNAc III. These results ind icate that the expressed enzyme is a new type of GalNAc alpha 2,6-sialyltra nsferase, which requires sialic acid residues linked to Gal beta 1,3GalNAc residues for its activity; therefore, we designated it mouse ST6GalNAc TV. Although the substrate specificity of this enzyme is similar to that of ST6 GalNAc III, ST6GalNAc IV prefers O-glycans to glycolipids, Glycolipids, how ever, are better substrates for ST6GalNAc III.