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S Manzella

Publications and source records attributed to S Manzella.

7 recordsLinked to original sources
PubMed1997

Alkylglycosides as artificial primers for glycogen biosynthesis.

Glycogenin is a 37 kDa self-glycosylating protein which has been demonstrated to be the initiating enzyme and primer for glycogen biosynthesis in liver, skeletal muscle and other tissues. We have recently shown that glycogenin will use alkylglucosides and alkylmaltosides as artificial acceptors in glycosyl transfer from UDP-glucose and UDP-xylose in vitro and have suggested that such substrates might be used to promote the synthesis of glycogen in vitro and in vivo. We now report that alkylglycosides can also serve as acceptors for transfer of glucose by glycogen synthase, yielding alkylmaltooligosaccharide products which may potentially be elongated to glycogen. alpha-Glucosides were better substrates than the corresponding beta-glucosides, and alkylmaltosides were preferred over alkylglucosides. The hydrophobicity of the substrates markedly affected their acceptor activity, less hydrophobic substrates being more active. This is in contrast to the behavior of glycogenin, which acted preferentially upon the more hydrophobic substrates tested. Aromatic glycosides were also substrates for glycogen synthase, e.g., naphthyl-alpha-D- and beta-D-glucoside. The substrates were active in vitro both with partially purified rabbit muscle glycogen synthase and in incubations with crude muscle and liver homogenates from rat. In vivo experiments with mice further proved that intraperitoneal administration of alkylglucosides and alkylmaltosides increased the uptake of 14C-glucose in liver. The elevated uptake was due to an increase in both hydrophobic products, isolated by adsorption to Sep-Pak C18 columns, and more hydrophilic material that co-fractionated with glycogen upon treatment of the tissue with alkali and precipitation with ethanol. These results demonstrate the ability of alkylglycosides to serve as artificial primers for glycogen biosynthesis in vivo.

Alkylation↗
PubMed1995

Inhibition of glycogenin-catalyzed glucosyl and xylosyl transfer by cytidine 5'-diphosphate and related compounds.

The self-glucosylation of beef kidney glycogenin was inhibited by the following pyrimidine nucleotides and nucleotide sugars, listed in order of decreasing effectiveness: CDP-glucose, CDP, UDP-xylose, UDP-N-acetylglucosamine, UDP-galactose, UDP, CTP, CDP-choline, UDP-glucuronic acid, beta-S-UDP-glucose, and CMP. In contrast, the purine nucleotide sugars, ADP-glucose and GDP-glucose, were essentially ineffective, as was the pyrimidine nucleoside, cytidine. UDP-Xylose may be utilized by glycogenin as an alternative sugar donor instead of UDP-glucose (Rodén, L., Ananth, S., Campbell, P., Manzella, S., and Meezan, E. (1994) J. Biol. Chem. 269, 11509-11513) and therefore presumably inhibited the glucosyl transfer reaction by being a competitive substrate. Like glucosyl transfer, xylosyl incorporation into glycogenin was also inhibited effectively by CDP. On the other hand, UDP-xylose:proteoglycan core protein xylosyltransferase (EC 2.4.2.26) was not affected by CDP, nor was it inhibited by UDP-glucose. Addition of CDP or UDP-glucose to reaction mixtures containing both enzymes therefore made it possible to assay xylosyltransferase EC 2.4.2.26 reliably without the extensive product characterization that is otherwise necessary. The CDP effect on glycogenin further allowed the development of an improved procedure for the purification of this enzyme, in which specific elution of an affinity matrix (UDP-glucuronic acid-agarose) was carried out with CDP as the eluant.

Animals↗
PubMed1994

Xylosyl transfer to an endogenous renal acceptor. Characteristics of the reaction and properties of the product.

In the course of a study of UDP-xylose:proteoglycan core protein xylosyltransferase (EC 2.4.2.26), another xylosyltransferase was discovered in the soluble fraction of a rat kidney homogenate. The latter enzyme catalyzed [3H]xylosyl transfer from UDP-[3H]xylose to an endogenous acceptor and yielded a product in which the xylose was bound by an alkali-stable linkage. It was therefore concluded that the acceptor was not the core protein of one of the proteoglycans containing a xylose-->serine linkage, since this linkage is cleaved by alkali. The [3H]xylose-labeled product emerged with the void volume when chromatographed on Sephadex G-50, it was precipitated by trichloroacetic acid, and it had a mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis corresponding to a molecular mass of about 32,000 Da. Digestion with trypsin or alpha-amylase degraded the labeled product to small fragments, as determined by gel chromatography, suggesting that it was a glycoprotein related to glycogen. A product of similar characteristics was formed when UDP-[3H]glucose was substituted for UDP-[3H]xylose as the glycosyl donor, and the two nucleotide sugars were mutually competitive in the respective transfer reactions, indicating that they were substrates for the same enzyme. On the basis of these findings, it was tentatively concluded that the xylosyltransferase and its acceptor were the renal form of glycogenin.

Animals↗
PubMed1994

Xylosyl transfer to an endogenous renal acceptor. Purification of the transferase and the acceptor and their identification as glycogenin.

A xylosyltransferase in rat kidney, tentatively identified as glycogenin (Meezan, E., Ananth, S., Manzella, S., Campbell, P., Siegal, S., Pillion, D. J., and Rodén, L. (1994) J. Biol. Chem. 269, 11503-11508), was purified by a procedure in which affinity chromatography on UDP-glucuronic acid-agarose was a particularly useful step. The purified material was nearly homogeneous, as shown by SDS-polyacrylamide gel electrophoresis and silver staining, and had an electrophoretic mobility corresponding to a M(r) of 32,000. The purified enzyme possessed both glucosyl- and xylosyltransferase activity, and incubation with UDP-[3H]xylose or UDP-[3H]glucose yielded a single macromolecular product, which had the same electrophoretic mobility as the major silver-stained component. These results indicate that the kidney transferase was indeed glycogenin and that it was functionally analogous to the larger glycogenin species previously isolated from rabbit muscle. Further examination of the properties of the rat kidney enzyme showed, i.a., that it was inhibited strongly by cytidine 5'-diphosphate. This effect was used to advantage in an alternative purification procedure, which was applied to beef kidney and involved adsorption of the enzyme to UDP-glucuronic acid-agarose and subsequent elution with cytidine 5'-diphosphate. In contrast to glycogenin, glycogen synthase did not catalyze transfer from UDP-xylose, and it is suggested that the incorporation of xylose into glycogen observed by other investigators was due to glycogenin-catalyzed xylosyl transfer and subsequent chain elongation by glycogen synthase.

Animals↗