Differentiated structure of synthetic glycogen-like particle by the combined action of glycogen branching enzymes and amylosucrase.

Glycogen-like particles (GLPs) were built up from sucrose by applying de novo one-pot enzymatic process of amylosucrase (ASase; 6 U·mL-1) and glycogen branching enzymes (GBEs; 0.001 and 0.005 U·mL-1). Due to different chain-length transferring patterns of GBEs, structurally differentiated GLPs were synthesized. Yields of GLPs synthesized at pH 7.0 and 30 °C were improved by increasing the GBE/ASase ratio. Branching degrees of GLPs obviously was increased along with the ratio of GBEs, of which result was directly supported by shortened branch-chain length with greater GBE activity. Long branch chains seemed to play as efficient acceptor molecules to bind newly transferred branch chains especially at lower ratio of GBE/ASase, resulting in greater molecular weight and size of GLP with higher proportion of them. Molecular weight, size, and density of GLPs were ranged from 7.37 × 105 to 1.94 × 108 g·mol-1, from 23.70 to 52.65 nm, and from 7.99 to 374.32 g·mol-1·nm-3, respectively. By increasing GBE/ASase ratio, more compact GLP architecture was fabricated due to increased weight and reduced size with exception of a unique GBE. GLPs were efficiently synthesized by two different glycosyltransferases, and their chemical structures were controllable by source and ratio of GBEs due to their different branch-chain transferring specificity.

[Dynamics of pro- and macroglycogen content in hepatocytes of normal and cirrhotic rat liver at different stages of glycogenesis].

The content and structure of glycogen in hepatocytes of normal and cirrhotic rat liver has been studied at definite time intervals after the administration of glucose to starving animals. In the study, an original cytofluorimetric method for detection and quatification of proglycogen (PG) and macroglycogen (MG) content in isolated hepatocytes was applied. This method is based on using Schiff reagents with different spectral characteristics. It has been determined that the content MG content in the hepatocytes of control rats increases in 10 min after initiation of glycogenesis by 52% (P < 0.01). MG content in the cells of cirrhotic liver increased only after 20 min (43%, P < 0.05) after glucose administration to starving animals. The coefficient of correlation between MG content and the total glycogen content in the hepatocytes at different stages of glycogenesis ranged from 0.90 to 0.99 (P < 0.001) in both groups of rats. Increase in PG content in hepatocytes of control rats appeared within 10-30 and 45-70 min. In the case of cirrhosis PG content increased only 60 min after the start of glycogenesis, but after 120 min it was 1.5 times higher than the control values (P < 0.001). The correlation coefficient between the PG and the total glycogen content in rat liver cells averaged 0.86 (P < 0.001) and 0.77 (P < 0.001) in control and experimental groups, respectively. Thus, the change in total glycogen content in hepatocytes of normal and cirrhotic liver is associated mainly with the level of MG. In normal cells, contribution of PG is most significant in the early glycogenesis (10-30 min), and in the cirrhotic liver--in the later stages.

The exchange between proglycogen and macroglycogen and the metabolic role of the protein-rich glycogen in rat skeletal muscle.

The aim of this study is to determine if proglycogen and macroglycogen are kinetically related in rat skeletal muscle. Eight groups of anesthetized fasted rats (seven hepatic-occluded and one nonoccluded) were intravenously infused with [3-3H]glucose at a rate of 1.7 microCi x min(-1) for 20 min. At the end of infusion, hindlimb muscles were excised and rapidly frozen in liquid nitrogen. Proglycogen was extracted by precipitation in 10% TCA; and macroglycogen as a part of total glycogen by precipitation in 20% KOH-65% ethanol. Along with the tracer, the occluded rats were also infused with: saline (group 1); insulin at rates ranging from 5 to 50 mU x min(-1) (groups 2 to 5); and insulin at a rate of 10 mU x min(-1) plus glucose at rates of 10.2 and 20.4 micromol x min(-1), respectively (groups 6 and 7). The infusion regimens resulted in up to 30-fold difference in whole-body glucose utilization among the rats. In the rats infused with saline and insulin at a rate of 5 mU x min(-1), [3H]glucose was found to be exclusively incorporated into proglycogen. Incorporation into macroglycogen was found in the rats infused with insulin at rates > 10 mU x min(-1). Supplementary glucose infusion increased the synthesis of [3H]proglycogen (four- to sixfold), and equilibrated the two extractable forms of glycogen in the insulin-infused rats. In the saline-infused nonoccluded rats, only proglycogen was found to be labeled. In conclusion, our data indicate that in the intact and hepatic-occluded rats, proglycogen in the skeletal muscles may undergo synthesis and degradation of its own more readily than exchange between itself and depot macroglycogen.