SUBSTRATE-SPECIFICITY AND INHIBITION OF UDP-GLCNAC-GLCNAC-BETA-1-2MAN-ALPHA-1-6R BETA-1-6-N-ACETYLGLUCOSAMINYLTRANSFERASE-V USING SYNTHETICSUBSTRATE-ANALOGS

UDP-GlcNAc:GlcNAc beta 1-2Man alpha 1-6R (GlcNAc to Man) beta 1,6-N-ac etylglucosaminyltransferase V (GlcNAc-T V) adds a GlcNAc beta 1-6 bran ch to bi- and triantennary N-glycans. An increase in this activity has been associated with cellular transformation, metastasis and differen tiation. We have used synthetic substrate analogues to study the subst rate specificity and inhibition of the partially purified enzyme from hamster kidney and of extracts from hen oviduct membranes and acute my eloid leukaemia leukocytes. All compounds with the minimum structure G lcNAc beta 1-2Man alpha 1 -6Glc/Man beta- were good substrates for Glc NAc-T V. The presence of structural elements other than the minimum tr isaccharide structure affected GlcNAc-T V activity without being an ab solute requirement for activity. Substrates with a biantennary structu re were preferred over linear fragments of biantennary structures. Kin etic analysis showed that the 3-hydroxyl of the Man alpha 1-3 residue and the 4-hydroxyl of the Man beta- residue of the Man alpha 1-6(Man a lpha 1-3)Man beta-R N-glycan core are not essential for catalysis but influence substrate binding. GlcNAc beta 1-2(4,6-di-O-methyl-)Man alph a 1- 6Glc beta-pnp was found to be an inhibitor of GlcNAc-T V from ham ster kidney, hen oviduct microsomes and acute and chronic myeloid leuk aemia leukocytes.