Retaining Glycosyltransferases

An enzyme which transfers the a-Gal residue to A-acetyllactosamine structures in the glycoproteins of mammals other than humans has attracted attention because the a-Gal/)-(1 3)-Gal epitope is a major obstacle to the use of organs from other mammals in xenotransplantation because of the 

The suggested nucleophile (likewise Glu 317) in the bovine enzyme has been mutated to Ala. 

The mutant enzyme has no transferase activity (although it has hydrolase activity) and can be catalytically rescued by azide ion the product from UDPGal is p-galactopyranosyl azide. This incisive experiment is powerful evidence for the double displacement, rather than S i, mechanism for this family.  

Although the enzyme aeeepted a-galaetosyl fluoride as a glycosyl donor, it did this only in the presenee of UDP UDPGal was first synthesised. Galaetosyl fluoride eould aet both as an aeceptor and an preeursor of UDPGal, so that if 

In the presence of the incompetent acceptor 4 -deoxylactose, there is no positional isotope exchange with UDPGal, nor is there any sign of enzyme inactivation by 5-fluoro-ot-galactosyl fluoride, even at high concentrations and in the presence of UDP. 

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Boix E, Zhang Y, Swaminathan GJ, Brew K, Acharya KR. Structural basis of ordered binding of donor and acceptor substrates to the retaining glycosyltransferase, a-l,3-galactosyltransferase. J. Biol. Chem. 2002 277 28310-28318. 

Scheme 6. Hypothetical two-step, double-displacement mechanism for retaining glycosyltransferases.

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