SYNTHESIS OF PENTASACCHARIDE ANALOGS OF THE N-GLYCAN SUBSTRATES OF N-ACETYLGLUCOSAMINYLTRANSFERASE-III, N-ACETYLGLUCOSAMINYLTRANSFERASE-IV AND N-ACETYLGLUCOSAMINYLTRANSFERASE-V USING TETRASACCHARIDE PRECURSORSAND RECOMBINANT BETA-(1-]2)-N-ACETYLGLUCOSAMINYLTRANSFERASE-II

Recombinant human UDP-GlcNAc:alpha-Man-(1 --> 6)R beta-(1 --> 2)-N-ace tylglucosaminyltransferase II (EC 2.4.1.143, GlcNAc-T II) was produced in the Sf9 insect cell/baculovirus expression system as a fusion prot ein with a (His)(6) tag and partially purified by affinity chromatogra phy on a metal chelating column. The partially purified enzyme was use d to catalyze the transfer of GlcNAc from UDP-GlcNAc to R-alpha-Man(1 --> 6)(beta-GlcNAc(1 --> 2)alpha-Man(1 --> 3))beta-Man-O-octyl to form beta-GlcNAc(1 --> 2)R-alpha-Man(1 --> 6)(beta-GlcNAc(1 --> 2)alpha Ma n(1 --> 3))beta-Man-O-octyl where there is either no modification of t he alpha-Man(1 --> 6) residue (7), or where R is 3-deoxy (8), 4-deoxy (9) or 6-deoxy (10). The yields ranged from 64-80%. Products were char acterized by H-1 and (13) C nuclear magnetic resonance spectroscopy an d fast atom bombardment mass spectrometry. Compounds 7-10 are pentasac charide analogues of the biantennary N-glycan substrates of N-acetylgl ucosaminyltransferases III, IV and V.