Glycolysis
4 / The Text :- Some Points About Glucose Glucose is very soluble source of quick and ready energy. It is a relatively stable and easily transported. In mammals, the brain uses only glucose under non-starvation conditions. Under starvation conditions ketone bodies become a source of energy for the brain Glucose is the only source of energy in red blood cells.
Cellular glucose transport Glucose gradient from extracellular to intracellular environment Glucose - polar molecule Membrane transporter required GLUT transporter family GLUT transporters in RBC, brain, liver and kidney cells unregulated GLUT transporters in skeletal muscle, fat and heart regulated Predominantly GLUT-4
Regulation of Cellular Glucose Uptake Brain & RBC: GLUT-1 has high affinity (low Km)for glucose and are always saturated. Insures that brain and RBC always have glucose. Liver: GLUT-2 has low affinity (hi Km) and high capacity. Uses glucose when fed at rate proportional to glucose concentration Muscle & Adipose: GLUT-4 is sensitive to insulin
Glycolysis Basic overview of glycolysis: Glycolysis converts glucose to two C3 units (pyruvate): Ten enzymes catalyse the reactions. First stage (5 reactions) in which glucose phosphorylated to yield two molecules of a triose: glyceraldehyde 3-phosphate. * Two molecules of ATP consumed. - Second stage (5 reactions) converts this triose to pyruvate. * Four molecules of ATP produced. - Net result is to produce two ATP and two NADH. Occurs in cytosol Under aerobic conditions, NADH can be converted to 2-3 ATPs
Glycolysis - Step 1 Glucose G-6-P Catalysed by hexokinase in skeletal muscle (glucokinase in liver) Non-equilibrium reaction Energy from ATP used to phosphorylate C6 Traps glucose in cell
glucose Hexokinase
Glycolysis - Step 1 Glucokinase has higher Km for glucose than Hexokinase Ensures hexokinase has first call on circulating glucose Glucokinase provides G-6-P for glycogen synthesis Not allosterically inhibited by G- 6-P Hexokinase Provides G-6-P for energy production Allosterically inhibited by G-6-P From: Mathews CK & van Holde KE (1990) Biochemistry. Redwood City: Benjamin Cummings p 439.
Glycolysis - Step 2 G-6-P Fructose-6-P Catalysed by Phosphoglucose isomerase Equilibrium reaction Generated hydroxyl group (-OH) at C1 Alters energy distribution within molecule Allows phosphorylation of C1 in next step
Glycolysis - Step 3 F-6-P Fructose 1,6-Bisphosphate Catalysed by Phosphofructokinase Non-equilibrium reaction First non-reversible reaction unique to glycolytic pathway Committed step Primary control site Inhibited by ATP, citrate and H + Stimulated by AMP, ADP, Epinephrine, Pi, NH 4 + Adds phosphate group to C1 prepares for cleavage into two phosphorylated molecules in next step
doubled from this point on Glycolysis - Step 4 Cleavage of F 1,6-Bisphosphate Catalysed by Aldolase Equilibrium reaction Cleaves 6C molecule of F 1,6- Bisphosphate to form two 3C molecules of DHAP and G-3-P G-3-P go on direct glycolytic pathway DHAP and G-3-P in equilibrium 96% held as DHAP All reactions in pathway
Glycolysis - Step 5 G-3-P 1,3-BPG Catalysed by G-3-P dehydrogenase Equilibrium reaction First reaction where energy produced (indirectly) G-3-P oxidised by removal of H NAD + reduced to NADH NADH to ETC to give 3 ATP Oxidation provides energy to incorporate Pi into structure as high energy phosphate group Doubling of reaction gives 2 x NADH = 6 ATP Oxidation of glyceraldehyde-3-phosphate to give 1,3-bisphosphoglycerate: The oxidizing agent, NAD+, is reduced to NADH: involves addition of a phosphate group, as well as an electron transfer
Glycolysis - Step 6 1,3-BPG 3-PG Catalysed by phosphoglycerate kinase Equilibrium reaction High energy phosphate group removed Energy release used to phosphorylate ADP to ATP (substrate level phosphorylation) Doubling of reaction gives 2 x ATP
Glycolysis - Step 7 3-PG 2-PG Catalysed by phosphoglyceromutase Equilibrium reaction Phosphate group moved from C3 to C2
2-PG PEP Glycolysis - Step 8 Catalysed by enolase Equilibrium reaction Dehydration causes redistribution of energy within molecule creating a high energy phosphate group
PEP Pyruvate Glycolysis - Step 9 Catalysed by Pyruvate kinase Non-equilibrium reaction High energy phosphate group removed Energy release used to phosphorylate ADP to ATP (substrate level phosphorylation) Intermediate step in reaction forms enolpyruvate which spontaneously converts to pyruvate Doubling of reaction yields 2 ATP
Lactic acid NADH produced at Step 5 reoxidised to NAD + in ETC If unable to oxidise NADH glycolysis would stop Energy only put in prior to step 5 Would be energy consuming pathway NADH usually oxidised in ETC NADH also oxidised by LDH reducing pyruvate to lactate
Lactic acid Lactate has numerous fates: Remain within cell until can be reoxidised to pyruvate Released from cell Taken up by other less active fibres in same muscle Enter circulation to be taken up by: other less active fibres in other skeletal muscle Heart for oxidation Liver for conversion to glucose via Cori cycle Release of lactate from skeletal muscle represents means of redistributing CHO stores throughout body
Glycolysis Enzyme/Reaction DG o ' kj/mol DG kj/mol Hexokinase -20.9-27.2 Phosphoglucose Isomerase +2.2-1.4 Phosphofructokinase -17.2-25.9 Aldolase +22.8-5.9 Triosephosphate Isomerase Glyceraldehyde-3-P Dehydrogenase & Phosphoglycerate Kinase +7.9 negative -16.7-1.1 Phosphoglycerate Mutase +4.7-0.6 Enolase -3.2-2.4 Pyruvate Kinase -23.0-13.9
Glycolysis: Energy- Production In reactions 7 and 10, the hydrolysis of phosphates in the triose phosphates generates four ATP molecules.
Glycolysis: Overall Reaction Glycolysis generates 2 ATP and 2 NADH. Two ATP are used in energy-investment to add phosphate groups to glucose and fructose-6-phosphate. Four ATP are formed in energy-generation by direct transfers of phosphate groups to four ADP. Glucose + 2ADP + 2P i + 2NAD + 2Pyruvate + 2ATP + 2NADH + 2H +
Regulation of Glycolysis Reaction 1 Hexokinase is inhibited by high levels of glucose-6-phosphate, which prevents the phosphorylation of glucose. Reaction 3 Phosphofructokinase, an allosteric enzyme, is inhibited by high levels of ATP and activated by high levels of ADP and AMP. Reaction 9 Pyruvate kinase, another allosteric enzyme is inhibited by high levels of ATP or acetyl CoA.
Control Points in Glycolysis Three reactions exhibit particularly large decreases in free energy; the enzymes that catalyze these reactions are sites of allosteric control Hexokinase Phosphofructokina se Pyruvate kinase
Hexokinase also Regulates Glycolysis Hexokinase is inhibited by its product, glucose 6-phosphate. High concentrations of glucose 6-phosphate indicates that the cell no longer needs glucose for energy, for storage as glycogen, or for other precursors. Remember that the liver is responsible for regulating blood glucose levels.
Hexokinase and Glucokinase. The liver contains an isoform of hexokinase called glucokinase. Glucokinase is not inhibited by glucose 6- phosphate. Glucokinase has a lower affinity for glucose than hexokinase. This assures that brain and muscle have first choice for the glucose. When glucose is abundant in the liver, glucokinase phosphorylates glucose to glucose 6-phosphate specifically for glycogen synthesis.
Pyruvate kinase has regulatory role in glycolysis Pyruvate Kinase has an L (liver) and M (muscle and brain) form. Both forms are inhibited by its product, ATP. Fructose 1,6-bisphosphate activates both forms of the enzyme to keep pace with the influx on intermediates. Alanine can be reversibly transaminated to pyruvate. Alanine also inhibits pyruvate kinase thus indicating that building blocks are abundant.
Only three enzymes function with large negative DG s Hexokinase, Phosphofructokinase and pyruvate kinase The other enzymes operate near equilibrium and their rates are faster than the flux through the pathway. Specific effectors of Glycolysis Enzymes Inhibitors Activators Hexokinase G6P none PFK ATP, citrate, PEP ADP, AMP, camp FBP,F2,6BP, F6P NH 4, Pi Pyruvate kinase ATP none
Pyruvate is a branching point Pyruvate O 2 O 2 fermentation Kreb s cycle mitochondria
Pathways for Pyruvate When oxygen is present in the cell, (aerobic conditions), pyruvate from glycolysis is decarboxylated to produce acetyl CoA and CO 2. O CH 3 C COOH + HS CoA + NAD + pyruvic acid O CH 3 C S CoA + CO 2 + NADH + H + acetyl CoA
Lactate Formation When oxygen is not available (anaerobic conditions), pyruvate is reduced to lactate, which replenishes NAD + to continue glycolysis. O lactate dehydrogenase CH 3 C COO - + NADH + H + pyruvate OH CH 3 CH COO - + NAD + lactate
Lactate in Muscles Under anaerobic conditions (strenuous exercise): Oxygen in the muscles is depleted. Lactate accumulates in the muscles. Muscles tire and become painful. Rest is needed to repay the oxygen debt and to reform pyruvate in the liver.
Phosphorylation of Pyruvate Kinase When blood-glucose levels are low the glucagon camp cascade phosphorylates pyruvate kinase making it less active. This covalent regulation assures that brain and muscle get glucose when needed. Pyruvate kinase: only L-form is regulated by covalent modification.
Pathways for Pyruvate
Transition of pyruvate to acetyl CoA
Hexokinase also Regulates Glycolysis Hexokinase is inhibited by its product, glucose 6-phosphate. High concentrations of glucose 6-phosphate indicates that the cell no longer needs glucose for energy, for storage as glycogen, or for other precursors. Remember that the liver is responsible for regulating blood glucose levels.
Hexokinase and Glucokinase. The liver contains an isoform of hexokinase called glucokinase. Glucokinase is not inhibited by glucose 6- phosphate. Glucokinase has a lower affinity for glucose than hexokinase. This assures that brain and muscle have first choice for the glucose. When glucose is abundant in the liver, glucokinase phosphorylates glucose to glucose 6-phosphate specifically for glycogen synthesis.
Control Points in Glycolysis Reaction 3 Phosphofructokinase, an allosteric enzyme, is inhibited by high levels of ATP and activated by high levels of ADP and AMP. Reaction 9 Pyruvate kinase, another allosteric enzyme is inhibited by high levels of ATP or acetyl CoA.
The Overall Pathway of Glycolysis Glycolysis is the first stage of glucose metabolism One molecule of glucose is converted to fructose-1,6- bisphosphate, which gives rise to two molecules of pyruvate It plays a key role in the way organisms extract energy from nutrients 2 ATP molecules forms at the end of conversion of one glucose to two pyruvate. Conversion of glucose to pyruvate is an oxidation reaction which accompanied by reduction of NAD+ to NADH Once pyruvate is formed, it has one of several fates
5/ Post test Circle the correct answer : Which hormones are decrease is fasting State : ACTH. GH. Insulin. Hydrocortisone. There is no digestion and metabolism of CHO take place in he following : Mouth. Small intestine. Stomach, Large intestine. Fructose absorbed in small intestine and transport by : Active transport. Passively transport Active transport with carrier (Na). Active transport with carrier (K). The glucose is converted by kreb cycle to : Pyruvate. Lactate. Keton bodies. H2O+CO2+E. Dietary (CHO) are readily converted to glucose by : Digestion. Synthesis in liver Formed from glycerol.
Learning Check M1 Match the following with the terms below: (1) Catabolic reactions (2) Coenzymes (3) Glycolysis (4) Lactate A. Produced during anaerobic conditions B. Reactions that convert glucose to pyruvate C. Metabolic reactions that break down large molecules to smaller molecules + energy D. Substances that remove or add H atoms in oxidation and reduction reactions
Solution M1 Match the following with the terms below: (1) Catabolic reactions (2) Coenzymes (3) Glycolysis (4) Lactate A. 4 Produced during anaerobic conditions B. 3 Reactions that convert glucose to pyruvate C. 1 Metabolic reactions that break down large molecules to smaller molecules + energy D. 2 Substances that remove or add H atoms in oxidation and reduction reactions
Content Review What is glycolysis? What is the net ATPs produced from one glucose molecule under aerobic conditions? Under anaerobic conditions? What is the end product of glycolysis? The citric acid cycle begins with What are some vitamins that are used in energy metabolism? What is lactate?