SYNTHESIS AND EVALUATION OF THIO-TRISACCHARIDES AS ACCEPTORS FOR N-ACETYLGLUCOSAMINYLTRANSFERASE-V

N-Acetylglucosaminyltransferase-V (GlcNAcT-V) transfers a beta-linked GlcNAc residue from UDP-GlcNAc to the 6-OH group of the alpha Man resi due in oligosaccharides terminating in the sequence lcpNAc-(1-->2)-alp ha-D-Manp-(1-->6)-beta-D-Glcp-OR (5, R = (CH2)(7)CH3). The terminal Gl cNAc moiety may be replaced by a Glc residue to produce trisaccharide 6. Two thio analogs (7, 8) of trisaccharide 6, where the oxygen atoms in the glycosidic linkages between sugar residues were replaced by sul fur, were prepared by multistep chemical synthesis that made use of th e key intermediates 1,2-anhydro-3,4,6-tri-O-benzyl-alpha-D-glucose (10 ) and 1,2-anhydro-3,4,6-tri-O-benzyl-beta-D-manno (13) as donors for t he glycosylations. The thio analogs (7, 8) were kinetically evaluated as substrates for GlcNAcT-V and found to be accepters with two- to thr ee-fold increase in V-max but higher K-m values (7, K-m = 376 mu M; 8, K-m = 300 mu M) than their parent compound 6 (K-m = 111 mu M), which has the natural oxygen linkage. The thio analogs 7 and 8 could be quan titatively converted into the expected product tetrasaccharides (27, 2 8) by incubation with GlcNAcT-V and UDP-GlcNAc. The enzymatic results indicate that GlcNAcT-V tolerates the substitution of the natural oxyg en linkage of the acceptor by a sulfur linkage.