Purification of rabbit skeletal muscle proteoglycogen: studies on the glucosyltransferase activity of polysaccharide-free and -bound glycogenin

• Published in: Glycobiology (Oxford) Vol. 7; no. 4; pp. 571 - 578
• Main Authors: Carrizo, Maria E., Miozzo, Maria C., Goldraij, Ariel, Curtino, Juan A.
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01.06.1997
• Subjects: Animals; Chemical Precipitation; Chromatography, Ion Exchange; Detergents; Glucosyltransferases - metabolism; Glycogen - metabolism; glycogenin; glycogenin autogalactosylation; Glycoproteins - chemistry; Glycoproteins - isolation & purification; Glycoproteins - metabolism; Glycosylation; Molecular Weight; Muscle Proteins - isolation & purification; Muscle Proteins - metabolism; Muscle, Skeletal - chemistry; Polysaccharides - metabolism; Potassium Iodide; proteoglycogen; proteoglycogen activity; proteoglycogen isolation; Rabbits; Solubility; Uridine Diphosphate Glucose - metabolism
• ISSN: 0959-6658; 1460-2423
• DOI: 10.1093/glycob/7.4.571

Proteoglycogen is the end product in the process of glycogen biogenesis. We have purified rabbit muscle proteoglycogen and studied the glucosyltransferase reactions catalyzed by its protein moiety, glycogenin, free or bound to the polysaccharide. The purification strategy involved dissolution of proteoglycogen and cosedimenting membrane vesicles in a Triton X-114/Triton X-45 mixture followed by partition in the aqueous phase, potassium iodide precipitation of accompanying proteins, and washing by high-speed centrifugation. Glycogenin or a proteoglycogen species of an average molecular mass of 200 kDa was isolated by ion-exchange chromatography after the purified proteoglycogen had been subjected to long or short amylolytic digestion, respectively. Besides autoglucosylation from UDP-glucose, glycogenin was capable of autogalactosylation from UDP-galactose. The autoglucosylation reaction was not inhibited by the simultaneous glucosylation of the exogenous acceptors N-(maltosyl-α-1-4-(1-deoxiglucitol))peptide or n-dodecyl-β-D-maltoside. The polysaccharidebound glycogenin species of 200 kDa showed to be active for the glucosylation of exogenous acceptor and represented the isolated proteoglycogen of higher size having glucosyl transferase activity. This is the first description of the isolation of native proteoglycogen and a proteoglycogen species having glucosyltransferase activity.