Galactan biosynthesis in snails: a comparative study of beta-(1 -> 6) galactosyltransferases from Helix pomatia and Biomphalaria glabrata

Adult snails synthesize in their albumen glands a polysaccharide which is c omposed exclusively of D- or D- and L-galactose (Gal) residues which are in terglycosidically linked by 1 --> 3 and 1 --> 6 bonds. It is the only carbo hydrate source for embryos and freshly hatched snails. Two galactosyltransf erases are described in this study which are most likely involved in the bi osynthesis of this polysaccharide. One identified in Helix pomatia acts on oligosaccharides and could be used to synthesize a tetrasaccharide when the branched trisaccharide D-Gal-beta-(1 --> 3)-[D-Gal beta-(1 --> 6)]-D-Gal b eta -1 --> OMe was offered as acceptor. This enzyme, requiring Mg++- and Mn ++-ions for activity, introduced a linear beta-(1 --> 6) linkage at the ter minal non-reducing ends and was not detected in Biomphalaria glabrata. The other enzyme, which introduced beta-(1 --> 6) linkages at subterminal D-Gal residues, thus forming branching points in the polysaccharide, was found i n H. pomatia Arianta arbustorum and B. glabrata with comparable activities. With the enzyme preparation of H. pomatia, up to four D-Gal residues were introduced into vicinal positional forming single-membered side chains, if a hexasaccharide with five linearly beta-(1 --> 3)-linked D-Gal residues wa s offered as a acceptor. The multiple-branched structure formed is typical for snail galactans, making this enzyme a prime candidate for the branching enzyme in galactan synthesis. The enzyme activity could be solubilized and purified by affinity chromatography. In SDS-polyacrylamide electrophoresis , the Helix-derived eluate displayed two bands (68, 37 kDa) and that of Bio mphalaria ia five bands (68, 63, 17.5; 15, 13 kDa). The purified material s howed only 8% of the total activity of the crude extracts, but it could be shown that a phosphatase present in the crude extract can degrade UDP forme d in the transfer reaction and thus drive the reaction to completion.