Molecular Cloning and Functional Expression of Two Members of Mouse NeuAcα2,3Galβ1,3GalNAc GalNAcα2,6-Sialyltransferase Family, ST6GalNAc III and IV

Abstract

Two cDNA clones encoding NeuAc␣2,3Gal␤1,3GalNAc GalNAc␣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 specificity of the expressed recombinant enzyme, while the other cDNA clone includes an open reading frame coding for 302 amino acids. The deduced amino acid sequence is not identical to those of other cloned mouse sialyltransferases, although it shows the highest sequence similarity with mouse ST6GalNAc III (43.0%). The expressed soluble recombinant enzyme exhibited activity toward NeuAc␣2, 3Gal␤1,3GalNAc, fetuin, and GM1b, while no significant activity was detected toward Gal␤1,3GalNAc or asialofetuin, or the other glycoprotein substrates tested. The sialidase sensitivity of the 14 C-sialylated residue of fetuin, which was sialylated by this enzyme with CMP-[ 14 C]NeuAc, was the same as that of ST6GalNAc III. These results indicate that the expressed enzyme is a new type of GalNAc␣2,6-sialyltransferase, which requires sialic acid residues linked to Gal␤1,3GalNAc residues for its activity; therefore, we designated it mouse ST6GalNAc IV. Although the substrate specificity of this enzyme is similar to that of ST6GalNAc III, ST6GalNAc IV prefers O-glycans to glycolipids. Glycolipids, however, are better substrates for ST6GalNAc III.Sialic acids are key determinants of carbohydrate structures that play important roles in a variety of biological functions, like cell-cell communication, cell-substrate interaction, adhesion, and protein targeting. The transfer of sialic acids from CMP-Sia 1 to the terminal positions of the carbohydrate groups of glycoproteins and glycolipids is catalyzed by a sialyltransferase. Although roles of sialic acids have been proposed in the regulation of many biological phenomena, the purpose of this structural diversity remains largely obscure. To determine the meaning of the diversity of and the regulatory mechanism for the sialylation of glycoconjugates, it is necessary to obtain information on the enzymes themselves and the gene structure of sialyltransferases. Each sialyltransferase exhibits strict specificity for acceptor substrates and linkages (3-6). Although three linkages, Sia␣2,6Gal, Sia␣2,3Gal, and Sia␣2,6GalNAc, are commonly found in glycoproteins (7), and two, Sia␣2,3Gal and Sia␣2,8Sia, occur frequently in gangliosides (8), each of these linkages has been found in both gangliosides and glycoproteins (8 -10).So far, the cloning of three members of the ␣2,6-sialyltransferase family (ST6GalNAc I, II and III) has been reported (11-14). The cDNAs of ST6GalNAc I and II were cloned from both chick (11, 12) and mouse (13,62).2 The overall amino acid sequence identity of chick ST6GalNAc I is 30.5% to chick ST6GalNAc II, 43.2% to mouse ST6GalNAc I, and 33.6% to mouse ST6GalNAc II, and that of mouse ST6GalNAc I is 29.6% to mouse ST6GalNAc II and 28.3% to chick ST6GalNAc II, and that of chick ST6GalNAc II is 57.3% ...

Synthesis of Disialyl Lewis a (Lea) Structure in Colon Cancer Cell Lines by a Sialyltransferase, ST6GalNAc VI, Responsible for the Synthesis of α-Series Gangliosides

“…ST6GalNAc III was poorly active for a type I structure (14), and a human homolog has not yet been reported. ST6GalNAc IV was a protein-dominant (O-glycan) enzyme (14). Consequently, we concentrated our efforts on the function of ST6GalNAc V and VI.…”

“…Nomenclature of Cloned Glycosyltransferase-Six members of the GalNAc ␣2,6-sialyltransferase (ST6GalNAc) subfamily have been cloned so far: ST6GalNAc I (12), ST6GalNAc II (13), ST6GalNAc III (14), ST6GalNAc IV (14), ST6GalNAc V (10), and ST6GalNAc VI (11).…”

Molecular Cloning of Brain-specific GD1α Synthase (ST6GalNAc V) Containing CAG/Glutamine Repeats

“…Nomenclature of Cloned Sialyltransferase-Four members of the GalNAc ␣2,6-sialyltransferase (ST6GalNAc) subfamily have been cloned to date: ST6GalNAc I (12), ST6GalNAc II (13,14), ST6GalNAc III (10,11), and ST6GalNAc IV (11). The GalNAc ␣2,6-sialyltransferase cloned in this study is referred to as ST6GalNAc V according to Tsuji et al (15).…”

“…Although several ST6GalNAc cDNAs that may synthesize GD1␣ have been reported (10,11), ST6GalNAc V is specific for GM1b in contrast with other members of the ST6GalNAc family. Moreover, ST6GalNAc V showed brain-specific expression, suggesting a critical role of ST6GalNAc V in the synthesis of GD1␣ in brain tissues.…”

Functional Diversity of Mammalian Sialyltransferases

“…H o w e v e r, h u m a n S T 6 G a l N A c -V I c a n u t i l i z e f e t u i n as an acceptor substrate (105). The oligosaccharide NeuAcα2,3Galβ1,3GalNAc is a preferred substrate for ST6GalNAc-IV, but not for ST6GalNAc-III (97,99). The ganglioside GM1b is a preferred substrate for ST6GalNAc-III, -V, and -VI, but not for ST6GalNAc-IV (96-99, 101 -103) (Fig.…”