During the purification of the enzyme glycogen synthase from the muscle of the nematode Ascaris suum, approximately 70% of the glycogen synthase activity can be separated from the bulk of cellular glycogen by centrifugation for 60 min at 105,000 x . The glycogen synthase in the supernatant fraction has an Mr of 1.2 x 106 as determined by Sepharose 4B gel filtration chromatography. The glycogen synthase in this high molecular weight complex (glycogen primer complex) can be further purified by ConA-Sepharose affinity chromatography; the enzyme activity was eluted with 100 .mM a-methylmannoside. The glycogen synthase in glycogen'primer complex is predominately in the glucose 6-phosphatedependent form. The glycogen primer complex can catalyze the transfer of glucosyl units from UDP-glucose to an endogenous acceptor in the absence of exogenous glycogen. Analysis by SDS-PAGE showed three proteins (Mr 140,000, 78,000 and 34,000) and a carbohydrate polymer. The carbohydrate polymer can be partially digested with a-amylase. The glycogen primer complex was further digested by acid hydrolysis, and upon descending paper chromatography analysis, eight different carbohydrates were isolated, two of which were tentatively identified as glucose and sialic acid. The [14 C]-autoradiograph showed that in vitro synthesis of a glycogen-like polysaccharide occurred on this carbohydrate polymer. Polyclonal antibodies have been made to the glycogen primer complex, and Western Blot analysis indicated that all three proteins of the glycogen primer complex were antigenic. Collectively, the data indicate that a glycogen-like polysaccharide is synthesized from a carbohydrate-associated protein primer in the muscle of this worm.
| Identifer | oai:union.ndltd.org:unt.edu/info:ark/67531/metadc935639 |
| Date | 08 1900 |
| Creators | Ghosh, Paritosh |
| Contributors | Donahue, Manus J., Masaracchia, Ruthann A., Uphouse, Linda, O'Donovan, Gerard A., Kester, Andrew S. |
| Publisher | North Texas State University |
| Source Sets | University of North Texas |
| Language | English |
| Detected Language | English |
| Type | Thesis or Dissertation |
| Format | vii, 86 leaves : col. ill., Text |
| Rights | Public, Ghosh, Paritosh, Copyright, Copyright is held by the author, unless otherwise noted. All rights |
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