Studies On The Molecular Structure Of Animal Liver Glycogen And The Chain Stoppage Mechanism In Glycogen Molecule Growth

Glycogen is the key energy storage in animals including human and other eukaryotes. Glycogen is widely distributed in body, it mainly located in liver and skeletal muscle, the regular metabolism of liver glycogen is of great significance for maintaining the blood sugar under control. Glycogen is a highly branched glucose polymer, its structure can be classified in three structural levels:(1) the most basic structure is the chain-length distribution (CLD), which is the relative number of chain, linked by α-(1,4)-glycosidic linkages, as a function of DP. These chains are joined together via α-(1,6)-glycosidic linkages to give (2) highly branched β particles which are ～20 nm in diameter. The beta-particles are joined together to give (3)α particles(-100 nm in diameter) which show a cauliflower-like appearance in transmission electron microscopy(TEM).In our previous study, we utilized DMSO SEC to analyse the molecular structure of glycogen from db/db mice and healthy mice. It is shown that db/db mice have an impaired ability to synthesize the large composite glycogen a particles present in normal. This discovery has important potential implications for diabetes management, because there is evidence that the rate of degradation per monomer unit of glycogen to glucose wille be faster in samller β particles.healthy mice and that a particles are held together via a bond more acid-labile than normal glycosidic linkages, with the most likely bond being proteinaceous. The structure of healthy mouse-liver glycogen over the diurnal cycle is characterized using size exclusion chromatography (SEC) and TEM. Glycogen is observed to be initially formed as smaller β particles, only being assembled into the larger α particles significantly when glycogen is in degradation state. This pathway which is impaired in db/db mice is likely to give the optimal blood glucogen control. It has been proved an aqueous-SEC system has a better separation/resolution of a particles and P particles in glycogen. Thus this thesis is to deeply study on animal liver glycogen structure and the chain stoppage mechanism in glycogen molecule growth.This thesis is divided in four parts.The first part:In this part we utilize water SEC to analyse the molecular structure of glycogen from db/db mice and healthy mice and simulate glycogen degradation in vitro. The water SEC results show that liver glycogen from db/db mice also have a big amount of a particles like glycogen from healthy mice. However, it is found that a particles from db/db mice liver glycogen is vulnerable to be degraded into samller 0 particles in DMSO and a particles from healthy mice liver glycogen is stable in DMSO. The results of glycogen degradation in vitro show that the degradation rate of β particles is bigger than a particles. The Km of glycogen phosphorylase to glycogen form db/db mice and healthy mice is 11.88±0.92 mM and 41.93±3.43 mM respectively. A smaller Km means glycogen phosphorylase is easier to collect with glycogen from db/db mice. All the data show that the formation of a particles is impared in db/db mice and the unstable a particles might contribute the high blood suger level in db/db mice.The second part:The present study reports the first molecular structural characterization of human-liver glycogen from non-diabetic patients, using TEM for morphology, fluorophore-assisted carbohydrate electrophoresis (FACE) for CLD and SEC for the molecular size distribution; the latter is also studied as a function of time during acid hydrolysis in vitro, which is sensitive to certain structural features, particularly glycosidic vs. proteinaceous linkages. The CLD data show that human liver glycogen have a higher content of long chains than pig and mice liver glycogen. The size distribution result shows that fasted human liver glycogen only contain a particles. Although the reason of the absence of β particles is unknown, the aggregation state, size and molecular weight of a particles is similar to those in mice and pig liver glycogen. The hydrolysis results are compared with those seen in mice and pigs. The molecular structural change during acid hydrolysis is similar in each case, and indicates that the linkage of β into a particles is not glycosidic. This result, and the similar morphology in each case, together imply that human liver glycogen has similar molecular structure to those of mice and pigs.The third part:The mechanisms stopping the growth of individual chains in complex highly branched polymers are investigated by taking advantage of special properties of glycogen, a complex branched polymer of glucose. The CLD of individual chains of mice and humans, two very different mammalian species, show a component which is consistent with stoppage of the growth of individual chains by crowding (steric hindrance) as the molecule grows larger. This fits simple first-order chain-stopping kinetics, a mechanism which may well apply to stoppage by hindrance in other complex branched polymers.The fourth part:This part summarizes the recent study progress of glycogen on three aspects:glycogen structure, glycogen synthesis and degradation and glycogen related disease. Hope it will give some reference to the further study of glycogen.