Human liver glycogen phosphorylase inhibitors bind at a new allosteric site

AbstractBackground: Glycogen phosphorylases catalyze the breakdown of glycogen to glucose-1-phosphate for glycolysis. Maintaining control of blood glucose levels is critical in minimizing the debilitating effects of diabetes, making liver glycogen phosphorylase a potential therapeutic target.Results: The binding site in human liver glycogen phosphorylase (HLGP) for a class of promising antidiabetic agents was identified crystallographically. The site is novel and functions allosterically by stabilizing the inactive conformation of HLGP. The initial view of the complex revealed key structural information and inspired the design of a new class of inhibitors which bind with nanomolar affinity and whose crystal structure is also described.Conclusions: We have identified the binding site of a new class of allosteric HLGP inhibitors. The crystal structure revealed the details of inhibitor binding, led to the design of a new class of compounds, and should accelerate efforts to develop therapeutically relevant molecules for the treatment of diabetes

Modeling the Mechanism of Action of a DGAT1 Inhibitor Using a Causal Reasoning Platform

Triglyceride accumulation is associated with obesity and type 2 diabetes. Genetic disruption of diacylglycerol acyltransferase 1 (DGAT1), which catalyzes the final reaction of triglyceride synthesis, confers dramatic resistance to high-fat diet induced obesity. Hence, DGAT1 is considered a potential therapeutic target for treating obesity and related metabolic disorders. However, the molecular events shaping the mechanism of action of DGAT1 pharmacological inhibition have not been fully explored yet. Here, we investigate the metabolic molecular mechanisms induced in response to pharmacological inhibition of DGAT1 using a recently developed computational systems biology approach, the Causal Reasoning Engine (CRE). The CRE algorithm utilizes microarray transcriptomic data and causal statements derived from the biomedical literature to infer upstream molecular events driving these transcriptional changes. The inferred upstream events (also called hypotheses) are aggregated into biological models using a set of analytical tools that allow for evaluation and integration of the hypotheses in context of their supporting evidence. In comparison to gene ontology enrichment analysis which pointed to high-level changes in metabolic processes, the CRE results provide detailed molecular hypotheses to explain the measured transcriptional changes. CRE analysis of gene expression changes in high fat habituated rats treated with a potent and selective DGAT1 inhibitor demonstrate that the majority of transcriptomic changes support a metabolic network indicative of reversal of high fat diet effects that includes a number of molecular hypotheses such as PPARG, HNF4A and SREBPs. Finally, the CRE-generated molecular hypotheses from DGAT1 inhibitor treated rats were found to capture the major molecular characteristics of DGAT1 deficient mice, supporting a phenotype of decreased lipid and increased insulin sensitivity

Polyomic profiling reveals significant hepatic metabolic alterations in glucagon-receptor (GCGR) knockout mice: implications on anti-glucagon therapies for diabetes

<p>Abstract</p> <p>Background</p> <p>Glucagon is an important hormone in the regulation of glucose homeostasis, particularly in the maintenance of euglycemia and prevention of hypoglycemia. In type 2 Diabetes Mellitus (T2DM), glucagon levels are elevated in both the fasted and postprandial states, which contributes to inappropriate hyperglycemia through excessive hepatic glucose production. Efforts to discover and evaluate glucagon receptor antagonists for the treatment of T2DM have been ongoing for approximately two decades, with the challenge being to identify an agent with appropriate pharmaceutical properties and efficacy relative to potential side effects. We sought to determine the hepatic & systemic consequence of full glucagon receptor antagonism through the study of the glucagon receptor knock-out mouse (Gcgr^-/-) compared to wild-type littermates.</p> <p>Results</p> <p>Liver transcriptomics was performed using Affymetric expression array profiling, and liver proteomics was performed by iTRAQ global protein analysis. To complement the transcriptomic and proteomic analyses, we also conducted metabolite profiling (~200 analytes) using mass spectrometry in plasma. Overall, there was excellent concordance (R = 0.88) for changes associated with receptor knock-out between the transcript and protein analysis. Pathway analysis tools were used to map the metabolic processes in liver altered by glucagon receptor ablation, the most notable being significant down-regulation of gluconeogenesis, amino acid catabolism, and fatty acid oxidation processes, with significant up-regulation of glycolysis, fatty acid synthesis, and cholesterol biosynthetic processes. These changes at the level of the liver were manifested through an altered plasma metabolite profile in the receptor knock-out mice, e.g. decreased glucose and glucose-derived metabolites, and increased amino acids, cholesterol, and bile acid levels.</p> <p>Conclusions</p> <p>In sum, the results of this study suggest that the complete ablation of hepatic glucagon receptor function results in major metabolic alterations in the liver, which, while promoting improved glycemic control, may be associated with adverse lipid changes.</p

The effects of exercise during pregnancy upon maternal adipocyte characteristics and fetal growth in Wistar rats

This study examined the effects of maintaining exercise training throughout pregnancy upon metabolic and physical properties of parametrial fat cells and fetal growth in Wistar rats. Eight weeks prior to mating the animals (n=10) were trained to run for 2 hrs/day, 5 days/wk at 31 m/min up an 8 0 incline. Control animals (n=6) remained sedentary. All animals were mated and trained animals resumed running on the second day of gestation. There was no variation in body weight (p > 0.05) between the trained (T) and sedentary control (S) rats at mating but S weight was significantly greater (p 0.05). The rate of glucose oxidation (1C-1) by cells from T animals was significantly higher (p < 0.05) than the S in the presence of insulin, but much lower than oxidation rates of non-pregnant trained animals. In terms of the fetus, training reduced litter size (p < 0.05) and increased the incidence of fetal resorption. The results of this study indicate that the training adaptation of the adipocytes is largely lost during pregnancy but insulin responsiveness is maintained at a higher level than sedentary controls. This coupled with the apparent adverse effects of the training on the fetus suggests that exercise during pregnancy should be more closely investigated.Ball State UniversityMuncie, IN 47306Thesis (M.S.)--Ball State University, 1983

The effects of exercise during pregnancy upon maternal adipocyte characteristics and fetal growth in Wistar rats

This study examined the effects of maintaining exercise training throughout pregnancy upon metabolic and physical properties of parametrial fat cells and fetal growth in Wistar rats. Eight weeks prior to mating the animals (n=10) were trained to run for 2 hrs/day, 5 days/wk at 31 m/min up an 8 0 incline. Control animals (n=6) remained sedentary. All animals were mated and trained animals resumed running on the second day of gestation. There was no variation in body weight (p > 0.05) between the trained (T) and sedentary control (S) rats at mating but S weight was significantly greater (p 0.05). The rate of glucose oxidation (1C-1) by cells from T animals was significantly higher (p < 0.05) than the S in the presence of insulin, but much lower than oxidation rates of non-pregnant trained animals. In terms of the fetus, training reduced litter size (p < 0.05) and increased the incidence of fetal resorption. The results of this study indicate that the training adaptation of the adipocytes is largely lost during pregnancy but insulin responsiveness is maintained at a higher level than sedentary controls. This coupled with the apparent adverse effects of the training on the fetus suggests that exercise during pregnancy should be more closely investigated.Ball State UniversityMuncie, IN 47306Thesis (M.S.)--Ball State University, 1983

The hepatic PP1 glycogen-targeting subunit interaction with phosphorylase a can be blocked by C-terminal tyrosine deletion or an indole drug

AbstractThe inhibition of hepatic glycogen-associated protein phosphatase-1 (PP1-GL) by glycogen phosphorylase a prevents the dephosphorylation and activation of glycogen synthase, suppressing glycogen synthesis when glycogenolysis is activated. Here, we show that a peptide (280LGPYY284) comprising the last five amino acids of GL retains high-affinity interaction with phosphorylase a and that the two tyrosines play crucial roles. Tyr284 deletion abolishes binding of phosphorylase a to GL and replacement by phenylalanine is insufficient to restore high-affinity binding. We show that a phosphorylase inhibitor blocks the interaction of phosphorylase a with the GL C-terminus, suggesting that the latter interaction could be targeted to develop an anti-diabetic drug