Efficient preparation of natural and synthetic galactosides with a recombinant beta-1,4-galactosyltransferase-/UDP-4 '-gal epimerase fusion protein

The numerous biological roles of LacNAc-based oligosaccharides have led to an increased demand for these structures for biological studies. In this re port, an efficient route for the synthesis of beta -galactosides using a ba cterial beta -4-galactosyltransferase/-UDP-4 ' -gal-epimerase fusion protei n is described. The lgtB gene from Neisseria meningitidis and the galE gene from Streptococcus thermophilus were fused and cloned into an expression v ector pCW. The fusion protein transfers galactose to a variety of different glucose- and glucosamine-containing accepters, and utilizes either UDP-gal actose or UDP-glucose as donor substrates. A crude lysate from Escherichia coli expressing the fusion protein is demonstrated to be sufficient for the efficient preparation of galactosylated oligosaccharides from inexpensive UDP-glucose in a multigram scale. Lysates containing the fusion protein are also found to be useful in the production of more complex oligosaccharides in coupled reaction mixtures, e.g., in the preparation of sialosides from N-acetylglucosamine, Thus, bacterially expressed fusion protein is well sui ted for the facile and economic preparation of natural oligosaccharides and synthetic derivatives based on the lactosamine core.