Structures of Sugar Chains of a p-Nitrophenyl Acetate-Hydrolyzing Esterase from the Microsomes of Rat Liver1

The structures of sugar chains of a p-nitrophenyl acetate-hydrolyzing esterase from the microsomes of rat liver were established. The enzyme contained mannose and glucosamine as sugar components. Asparagine-linked sugar chains of the esterase were liberated by hydrazinolysis. After N-acetylation of the hydrazinolysate, the reducing ends of the sugar chains were coupled with 2-aminopyridine. Fluorescent pyridylamino (PA-) derivatives of sugar chains thus obtained were purified by gel filtration and reversed-phase HPLC. Eleven PA-sugar chains were obtained. The structures of the PA-sugar chains were first identified by HPLC using two series of separation systems by which 11 PA-oligomannose-type sugar chains with known structures could be separated. Further elucidation of the structures of each PA-sugar chain was performed by exoglycosidase digestions and partial acetolysis. The structures of two of the PA-sugar chains were further confirmed by 500 mHz 1H-NMR spectroscopy.