Human non-secretory ribonucleases. II. 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 P-4 chromatography and analysed by 600 MHz 1H-NMR spectroscopy and electrospray mass spectrometry. All four non-secretory RNase preparations contained the following structures: [formula: see text] 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.
Lawrence
C W
CW
Department of Chemistry, Lehigh University, Bethlehem, PA 18015.
Little
P A
PA
Little
B W
BW
Glushka
J
J
van Halbeek
H
H
Alhadeff
J A
JA
D001483:Base Sequence; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D007668:Kidney / Q000201:enzymology*; D008099:Liver / Q000201:enzymology*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009844:Oligosaccharides / Q000737:chemistry* / Q000302:isolation & purification; D011134:Polysaccharides / Q000737:chemistry* / Q000302:isolation & purification; D012260:Ribonucleases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D013154:Spleen / Q000201:enzymology*
D002241:Carbohydrates; D006023:Glycoproteins; D009844:Oligosaccharides; D011134:Polysaccharides; D012260:Ribonucleases