Open AccessIdentification of a Novel UDP‐GalNAc:GlcNAcβ‐ R β1–4 N ‐Acetylgalactosaminyltransferase from the Albumen Gland and Connective Tissue of the Snail Lymnaea stagnalis
Author(s) -
Mulder Hans,
Spronk Bertina A.,
Schachter Harry,
Neeleman Alex P.,
Eijnden Dirk H.,
JongBrink Marijke,
Kamerling Johannis P.,
Vliegenthart Johannes F. G.
Publication year - 1995
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1995.tb20374.x
Subject(s) - connective tissue , snail , biochemistry , enzyme , chemistry , glycoprotein , lymnaea stagnalis , biology , ecology , genetics
Both the albumen gland, one of the female accessory sex glands, and connective tissue of the freshwater snail Lymnaea stagnalis contain N ‐acetylgalactosaminyltransferase activity, capable of transferring GalNAc from UDP‐GalNAc in β1–4 linkage to the terminal GlcNAc residue of GlcNAcβ‐ R . The albumin gland enzyme was partially purified by affinity chromatography on UDP‐hexanolamine–Sepharose 4B. Using GlcNAcβ1–2Manα1–6(GlcNAcβ1–2Manα1–3)Manα1–4GlcNAcβ1–4GlcNAc or GlcNAcβ1–OMe as substrates, the enzyme showed an absolute requirement for Mn 2+ with an optimum concentration of 12.5–50 mM. The optimal pH was approximately pH 7.0. The enzyme activity was independent of the Triton X‐100 concentration in the range 0.25–2.5%, and no activation effect was found. The more labile connective tissue microsomal enzyme, subjected to the same optimization procedure, gave comparable results. Both enzyme activities have similar substrate specificities towards GlcNAc or GlcNAcβl–OMe, and towards oligosaccharides or glycopeptides with a non‐reducing terminal β‐GlcNAc unit, but cannot act on GlcNAcα1–OMe. Saccharides with non‐reducing terminal Gal or GalNAc residues, and free GalNAc, Gal or Glc residues are not acceptors. Product analysis was carried out for albumen gland N ‐acetylgalactosaminyltransferase and four acceptors having GlcNAcβ1‐ R as the terminal non‐reducing unit, and for connective tissue N ‐acetylgalactosaminyltransferase with GlcNAcβ1‐OMe as acceptor. In all instances, products with GalNAc β1–4‐linked to GlcNAc were obtained, showing that the connective tissue and the albumen gland activities are probably from one enzyme. This enzyme activity can be identified as UDP‐GalNAc:GlcNAcβ‐ R β1–4 N ‐acetylgalactosaminyltransferase, and is probably involved in the biosynthesis of N,N ′‐diacetyllactosediamine‐containing glycoproteins, like hemocyanin, in the snail L. stagnalis.