BIOCHEM 4.6 - GLUCONEOGENESIS & HORMONE SIGNALING (AN ONGOING LOOK AT GLUCOSE)

Glycolysis vs. Gluconeogenesis

• Glycolysis occurs in all cells to break down glucose for energy
• Gluconeogenesis occurs in the liver to maintain glucose homeostasis and ensure availability for red blood cells and the brain
• Regulation ensures glucose is broken down only when available, and glucose is built when stores are depleted

Overview of Gluconeogenesis

• Gluconeogenesis: De novo synthesis of glucose (reverse glycolysis)
• Process is slower than glycolysis
• Maintains blood glucose during fasting
• Occurs in the liver using precursors like lactate, amino acids, and glycerol
• There is no net gluconeogenesis from even-chain fatty acids but odd-chain and branched-chain can serve as minor precursors
• Even-chain fatty acids cannot serve as a carbon source for glucose because Acetyl-CoA cannot serve as a carbon source for glucose
• Oxidative decarboxylation of pyruvate into Acetyl-CoA is irreversible
• Two carbons are lost to CO2 during TCA cycle

Gluconeogenesis Enzymes

• Gluconeogenesis uses enzymes that overcome the irreversible stages of glycolysis
• Irreversible enzymes of glycolysis include:
• Glucokinase
• Phosphofructokinase-1
• Pyruvate kinase
• Gluconeogenesis enzymes include:
• Pyruvate carboxylase
• Phosphoenolpyruvate carboxylase
• Fructose-1,6-bisphosphatase
• Glucose-6-phosphatase

Gluconeogenesis from Lactate

• Involves both mitochondrial and cytosolic enzymes
• It is a relatively energy expensive process
• Lactate is converted back to glucose
• Lactate is converted to Pyruvate via Lactate Dehydrogenase (LDH)
• In mitochondrion, pyruvate is converted to oxaloacetate by pyruvate carboxylase (needs biotin and ATP)
• Oxaloacetate is reduced to malate via malate dehydrogenase
• Malate exits to cytosol and oxidized back to oxaloacetate by cytosolic malate dehydrogenase
• Oxaloacetate is decarboxylated to PEP via phosphoenolpyruvate carboxykinase (PEPCK) using GTP

Cori Cycle

• The Cori Cycle involves the lactate cycle in the liver, red blood cells, and muscle

Gluconeogenesis from Amino Acids and Glycerol

• Alanine and Glutamine are exported from muscle for gluconeogenesis
• They are the major source of carbon skeletons, but most other amino acids are also glucogenic
• These sources are converted into TCA cycle intermediates then to malate
• Alanine is converted to pyruvate via alanine aminotransferase
• Glycerol is the only component of fats that can be used for gluconeogenesis
• It enters at triose phosphates
• Steps include phosphorylation by glycerol kinase in liver
• Glycerol 3 phosphate dehydrogenase generates dihydroxyacetone phosphate (DHAP)
• Glycerol enters after pyruvate carboxylase (PC) and phosphoenolpyruvate carboxykinase (PEPCK) thus costing less ATP.

Regulation of Gluconeogenesis

• Primary regulation involves hormones
• Phosphorylation/dephosphorylation of enzymes are controlled by glucagon or insulin
• Primary control points are
• Phosphofructokinase-1
• Fructose-1,6-bisphosphase
• Fructose-2,6-bisphosphate is an allosteric effector that counter-regulates two opposing pathways and it activates PFK-1 and inhibits Fru-1,6-Bpase

Regulation of Gluconeogenesis with Fru-2,6-bisphosphate

• Fru-2,6-bisphosphate is produced by a bifunctional enzyme: PFK-2/Fru-2,6-Bpase
• It contains both kinase and phosphatase activity
• Glucagon leads to phosphorylation of fructose-2,6-bisphosphatase through protein kinase A
• Reduces Fru-2,6-BP decreasing stimulation of glycolysis at PFK-1
• Simultaneously reduces inhibition of gluconeogenesis, promoting a gluconeogenic mode in the liver
• Insulin mediates dephosphorylation thus activating PFK-2
• Increases Fru-2,6-BP activating PFK-1 and inhibits Fru-1,6-Bpase activity
• Triggers STORAGE rather than synthesis
• Gluconeogenesis is also regulated in mitochondria via Acetyl-CoA
• Increased hepatic Acetyl-CoA inhibits pyruvate dehydrogenase (PDH) and activates pyruvate carboxylase (PC)

Glucose Homeostasis

• Glucose homeostasis is maintained through hormonal regulation (Insulin and Glucagon)

Glucose Receptors

• GLUT2 involved in facilitated diffusion of glucose and fructose.
• GLUT2 Location: Liver, intestine, kidney, pancreatic β-cells, and brain
• Pancreatic islet β-cell uptake of glucose.
• High levels in the liver stimulate glycogenesis in an insulin-independent manner.
• In the fed state, responds to increased availability of glucose by increasing the rate of glucose uptake through the receptor
• Process stimulates insulin secretion.
• Intestine's function with it is to export glucose to plasma at the basolateral membrane.
• GLUT4 undertakes facilitated diffusion of glucose within skeletal and cardiac muscles, and adipose tissues
• It is insulin dependent
• Insulin signaling increases translocation of this receptor to the cell surface. Doubles the presence of the receptor.
• Muscle contraction increases this receptor expression, while fatty acids reduce translocation of this receptor to the cell surface.

Insulin and Insulin Signaling

• It is synthesized in the rough ER of pancreatic β-cells, and is transported via secretory vesicles from the Golgi
• Two polypeptide chains (alpha and beta) are joined by 2 disulfide bridges
• Preproinsulin is a single chain precursor that is cleaved into the signal peptide and proinsulin, and proinsulin is further split into C-peptide and insulin.
• Amino acid residues that bind the receptor are in blue boxes

Glucagon Signaling

• Stimulates glycogenolysis and gluconeogenesis
• Direct connection to the phosphorylation of the enzyme complex that regulates gluconeogenesis
• Triggers reductions in Fru-2,6-BP

Metabolic Effects of Insulin

• Promotes glucose uptake and glycogen synthesis in the liver and muscle
• Enhances glycolysis and lipogenesis in the liver
• Facilitates glucose uptake and triacylglycerol synthesis in adipose tissue

Metabolic Effects of Glucagon

• Inhibits glycolysis in the liver
• Stimulates glycogenolysis and gluconeogenesis in the liver

Glucagon and Liver

• Reaction cascade amplifies the hepatic response to glucagon

Epinephrine and Liver

• Activates glycogenolysis during stress to increase blood glucose
• Augments activities of glucagon on liver during severe hypoglycemia responses
• Phospholipase C
• Cleaves membrane phosphatidylinositol bisphosphate (PIP2) (phosphoprotein) leading to DAG and inositol trisphosphate (IP3, second messengers)
• Protein Kinase C
• Activated by DAG
• Serine-Threonine kinase

FED/Absorptive State

• Glycogen synthesis occurs in the liver and muscle
• Glycolysis is active in the liver and muscle
• Lipogenesis occurs in the liver
• Protein synthesis increases within muscle

Overnight Fasting State/Post-absorptive State

• Gluconeogenesis is active in the liver
• Lipolysis is increased in adipose tissue

Starvation State

• Gluconeogenesis is high in liver
• Ketogenesis is increased
• Lipolysis is high in adipose tissue

Regulation of Ketogenesis

• Ratio of Glucagon to Insulin determines fuel utilization and storage
• Insulin INHIBITS ketogenesis and stimulates acetyl CoA carboxylase
• High insulin occurs when glucose is converted to ATP and stored as Glycogen in Hepatocytes
• Fatty acids are converted to TAG and stored and synthesis of new molecules of fatty acids takes place for membranes and other needs
• Insulin levels decrease, and Glycogen stores are utilized for fuel
• Lipase activity increases and stored TAGs are released as Fatty Acids
• Fatty acids become major fuel source
• Malonyl-CoA is a regulator

Quick Summary

• Absorptive State
• Increased Citrate Lyase
• Increased Acetyl-CoA Carboxylase
• Increased Fatty Acid Synthase
• Active Lipoprotein Lipase in adipose capillary bed and inactive Lipoprotein Lipase at muscle unless exercise or activity is occurring
• Fasting/Starved State
• Decreased Elongation of fatty acids
• Utilization of fatty acids as a fuel source