Human Nonsecretory Ribonucleases .2. Structural Characterization Of The N-Glycans Of The Kidney, Liver And Spleen Enzymes By Nmr-Spectroscopy And Electrospray Mass-Spectrometry

The N-glycans have been removed by peptide-N-glycosidase F (PNGase F) from purified human non-secretory RNases derived from kidney, liver and spleen. The spleen RNase was purified by two procedures, one of which did not include the usual acid treatment step (0.25 M H2SO4, 45 min, 4-degrees-C), to determine if acid treatment alters the carbohydrate moieties. The N-glycans of the RNases were fractionated by Bio-Gel P4 chromatography and analysed by 600 MHz H-1-NMR spectroscopy and electrospray mass spectrometry. All four non-secretory RNase preparations contained the following structures:[GRAPHICS] The relative amounts of the trisaccharide, pentasaccharide and hexasaccharide appeared to vary slightly in the different tissue RNases. The overall results indicate: (i) that acid treatment during purification does not alter the N-glycans of non-secretory RNases; (ii) that the N-glycans from kidney, liver and spleen non-secretory RNases are very similar, if not identical, to one another, but different from the N-glycan structures reported for secretory RNase.