Gluconeogenesis

Role of gluconeogenesis in metabolism Synthesis of glucose from non carbohydrate sources. Requires (i) E from metabolism & (ii) source of carbons Essential for maintaining blood glucose concentrations Meets the bodys’ demands for glucose when carbohydrate stores are limited e.g. fasting & starvation Occurs primarily in the liver (90%) and < kidney (10%) Liver and Kidney have G-6-phosphatase activity Allows release of glucose into blood stream

Substrates for GNG Lactate (produced by anaerobic glycolysis e.g. in RBC’s and exercising skeletal muscle) Glucogenic aminoacids Glycerol TCA intermediates

Lactate can act as a substrate for GNG Lactate from exercising muscle diffuses into the blood stream In the liver lactate is converted to pyruvate by lactate dehydrogenase Produces NADH in the cytoplasm NADH

Glycerol from breakdown of Triglycerides can act as substrate for GNG ATP DHAP

Some amino acids can acts as substrates for GNG Amino acids can undergo transamination reactions - amino group transferred to -ketoglutarate End product is pyruvate or TCA intermediates

Alanine, cysteine, glycine, serine, threonine pyruvate Aspartate & asparganine oxaloacetate Phenylalanine & tyrosine fumarate Isoleucine, valine & methionine Succinyl Co A Arginine, glutamate, glutamine, histidine a-ketoglutarate Leucine, lysine, phenylalanine, tryptophan, tyrosine Acetoacetate and Acetyl Co A TCA No gluconeogenesis from fats

Pyruvate Glucose ’ aspartate & asparagine phenylalanine arginine, & tyrosine arginine, glutamate, glutamine, histidine isoleucine, valine, & methionine

Lactate & some aminio acids Some amino acids Glycerol

Gluconeogenesis is not the reversal of Glycolysis 2 pyruvate + 4 ATP + 2 GTP + 2 NADH + 2 H+ + 4 H20  glucose + 4 ADP + 2 GDP + 2 NAD+ + 6 Pi Glycolysis Glucose + 2 ADP + 2 Pi + 2 NAD+  pyruvate + 2 ATP + 2 H+ + 2 NADH + 2 H20

Bypass Reactions Pyruvate carboxylase & PEP carboxykinase bypass pyruvate kinase step Fructose 1,6 bisphosphatase bypasses Phosphofructokinase step 3. Glucose 6 phosphatase bypasses hexokinase step These provide for a spontaneous pathway in the direction of glucose synthesis -∆G is in the direction of sugar synthesis

1st bypass: Pyruvate is first converted to oxaloacetate Carboxylation reaction requiring E CO2 is added by Pyruvate carboxylase (mitochondrial enzyme) Most enzymes for GNG are cytoplasmic – only exception pyruvate + ATP + CO2 + H2O  oxaloacetate + ADP + Pi + 2H+

Oxaloacetate is shuttled into the cytosol and converted to (PEP) Oxaloacetate is synthesised in the mitochondria Oxaloacetate cannot diffuse out of the mitcohondria Converted to Malate and shuttled into the cytoplasm Uses a specific malate transport system

pyruvate oxaloacetate ATP CO2 ADP + Pi malate NADH NAD+ matrix cytosol phosphoenolpyruvate PEP carboxykinase Pyruvate carboxylase GTP GDP Malate dehydrogenase pyruvate(c)

2nd bypass: Fructose 1,6 bisphosphatase bypasses phosphofructokinase step F-6-P F-1,6-BP Pi Fructose 1, 6, bisphosphatase Phosphofructose kinase

3rd bypass: Glucose 6 phosphatase bypasses hexokinase step Glucose-6-P + H20  glucose + Pi Blood stream glucose Pi Glucose-6- phosphatase Hexokinase G-6-P

3rd bypass: Glucose 6 phosphatase bypasses the hexokinase step G-6-Pase is primarily an enzyme of liver (and kidneys) In hepatocytes the glucose-6-phosphatase reactions allows the liver to supply the blood with free glucose Muscle cells lack G-6-Pase and direct G-6-P to glycogen synthesis

G-6-P glucose Pi G-6-Pase Endoplasmic reticulum G-6-Pase is located on the membrane of the ER Hydrolysis of G-6-P releases glucose into the lumen of the ER Glucose is packaged into vesicles for transport

Reciprocal regulation of GNG and glycolysis Gluconeogenesis expends 6 P bonds of ATP and GTP 2 pyruvate + 4 ATP + 2 GTP + 2 NADH + 2 H+ + 4 H20  glucose + 4 ADP + 2 GDP + 2 NAD+ + 6 Pi Glycolysis yields 2 P bonds of ATP Glucose + 2 ADP + 2 Pi + 2 NAD+  pyruvate + 2 ATP + 2 H+ + 2 NADH + 2 H20 If two pathways runs concurrently becomes a futile cycle; must be regulated

Reciprocal regulation of GNG and glycolysis cont’d. When gluconeogenesis is on, glycolysis should be off When energy stores are high, glycolysis should be off When energy stores are low, glucose should be rapidly degraded to provide energy Regulation occurs at the sites of the irreversible reactions

Irreversible reactions provide regulation-1 Glucose-6-P + H2O  glucose + Pi G-6-P inhibits hexokinase (glycolysis) G-6-phosphatase activity (gluconeogenesis) is dependant on [G-6-P]

Irreversible reactions provide regulation – 2 cont’d F-6-P to F-1-6-BP : Phosphofructokinase (glycolysis) Enzyme Inhibited by ATP, citrate Stimulated by AMP F-1,6-BP to F-6-P : fructose 1,6-bisphosphatase (gluconeogenesis) Inhibited by AMP and stimulated by citrate.

Irreversible reactions provide regulation - 2 Fructose 2,6-bisphosphate reciprocally controls these two enzymes levels are controlled by glucagon and insulin levels are low during starvation – stimulates GNG levels are high during the fed state accelerates glycolysis.

Regulation of gluconeogenesis G-6-P Glucose enters liver after meal + - F-2,6-BP F-6-P PFK-2 P F-1,6-BPase Gluconeogenesis is inhibited X Insulin mediates dephosphorylation of PFK-2 PFK-1 F-1,6-BP PEP

Irreversible reactions provide regulation - 3 PEP to pyruvate : pyruvate kinase (Glycolysis) Enzyme inhibited by acetyl-CoA, ATP and alanine, signals that energy levels are high. Also controlled by phosphorylation by glucagon and insulin pyruvate kinase is inhibited during starvation

Irreversible reactions provide regulation – 3 cont’d Pyruavte to oxaloacetate: pyruavte carboxlyase (gluconeogeneis) stimulated by acetyl-CoA and inhibited by ADP more gluconeogenesis when energy levels are high. Phosphoenolpyruvate carboxykinase is similarly inhibited by ADP levels.