BIOL 541 1st Edition Lecture 13Outline of Last Lecture I. Metabolism Outline of Current Lecture II. GlycolysisIII. Thermodynamics and regulationCurrent LectureBiochem Lecture 13- Glycolysis: Reaction: The concentration of glucose inside the cell is zero, only glucose 6 phosphate is present.Phosphorylation of glucose:1. It traps glucose inside the cell as glucose is polar and can not cross the cell membrane except with the aid of glucose transporter. Glucose enters the cell and gets phosphorylated and can not leave until dephosphorylated.2. It maintains the concentration gradient thus enabling glucose to always enter the cell. Diffusion needs concentration gradient and difference in concentration gradient is always present as the glucose enters and gets phosphorylated.Reaction:1. C=O moves from the end to 2C because C=O are polar and pulls on electrons. This cleaves in the center enabling C=O to come closer to the bond due to strain and thus becomes easier to break.2. This in addition enables phosphorylation at the free end of 1C. Both the cleaved halves stay inside the cell. 3C with the center C=O is acetone (dihydroxy acetone) and hydrophobic so can not stay in cell. C1 is phosphorylated to form Fructose 1,6 bisphosphate. 2 ATP are consumed.3. Glyceraldehyde 3 phosphate gets oxidized to COOH, NADH acceptor. 2 NADH are formed : 1 from each half cleaved.4. Some oxidation and reduction reaction go through with a phosphate intermediate that ends ups as COOH attached to phosphate. Because COOH and phosphate or S is high energy bond, 2 ATP are formed. This is called as substrate level phosphorylation.These notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.5. H is deposited into the ETC (electron transport chain). In the end phosphate bond is energized toATP. Phosphate moves to central C and OH is removed to make water. Double bond fluctuates between central C and end C.6. Enol phosphate is energy compound that is used to make ATP and pyruvate. There is a bridge reaction where pyruvate is converted to acetyly COA for TCA cycle.Glycolysis:1 Glucose = 2 reduced NADH, 2+2 = 4-2= 2 ATP.Thermodynamics:Glycolysis and gluconeogenesis are the same except 3 irreversible steps.Free energy change:1. Phosphorylation of glucose is the free energy step. But isomerization of glucose 6 phosphate to fructose 6 phosphate is in equilibrium.2. Phosphorylation using fructose 6 kinase to form fructose 1,6 bisphosphate. This is the committed step. The other steps are in equilibrium.3. Phosphoenol pyruvate is converted to pyruvate. Lactic acid is regenerated for NAD.These 3 steps determine direction of substrate flow.Regulation:During fasting, you can call upon glucose reserves. Glucose stored as glycogen (polymer). Not a lot of it though only a few hours of supply. However, lots of fat present: so the peripheral tissues start consumingfat. Some organs like the brain can not use fat but need only glucose. Glucose comes from amino acid. Hence, gluconeogenesis is the process of making glucose from non carbohydrate sources.Fatty acids ----- 2 C units.Glucose needs atleast 3 C units.Peripheral tissues stimulate glycolysis for energy but inhibit gluconeogenesis. Liver inhibits glycolysis and stimulates gluconeogenesis as liver makes glucose for brain. Therefore, regulation depends on LOCATION.Peripheral tissues:(In the handout irreversible steps are displayed as bubbles).Glucose 6 phospho hexokinase is allosterically inhibited by glucose 6 phosphate- product inhibition.Glucose 6 phosphatase removes phosphate.Fructose 6 phosphate -- Phosphate -- ATP --- attach 1 C.Phosphofructokinase is used to make fructose 1,6 phosphate.Energy status of cell:If more energy is present, then this step is inhibited. But if low energy persists, glycolysis is stimulated viastimulation of phosphofructokinase.In the graph, ATP shifts the curve to the right and inhibits and decreases affinity for fructose 6 phosphate.Reverse reaction:Fructose 1,6 phosphatase is used to remove phosphate group from 1C to make fructose 6 phosphate.Pyruvate enters mitochondria where acetyl COA enters the TCA cycle. Acetyl COA is converted to acetyl citrate in TCA cycle.If citrate level increases, the citrate cycle process is inhibited and also glycolysis is inhibited.Citrate causes phospho fructose kinase to split as dimer (inactive) and tetramer (active). Product inhibition occurs downstream.Energy level:ATP -- ADPPhosphatase is inhibited by low energy.AMP stimulates phospho fructokinase.ADP stimulates ATP.Fructose 2,6 bisphosphate is made by phospho fructokinase 2/FBP2.1. Phospho fructo kinase phosphorylates 2C position.2. Phosphatase removes the phosphate group.3. The site decides the phosphorylation.4. The cAMP dependent kinase is needed for phosphorylation and activated by AMP.5. If energy is low and AMP is around, PFK2/FBP2 and convert phosphor fructokinase to synthesize fructose 2,6 bisphosphate which is the allosteric activator and inhibitor of PFK2/FBP2.6. If low levels of AMP exist then glycolysis is stimulated in tissues.Graph:Fructose 2,6 bisphosphate- curve shifts to right.Glucose 6 phosphate is 0.75 m.mol only and without it 2.0 m.mol. Therefore, 2 order of magnitude difference in affinity.Graph:Phosphatase stimulated in absence and curve shifts to right for inhibition. So, phosphorylated compoundis the regulator.Fructose 2,6 bisphosphate activates fructose phosphokinase and inhibits fructose 1,6 phosphatase.Aldolase splits phosphate compound into 2 hydroxyacetone and 2 glyceraldehyde.Phospho triose isomerase breaks 2 glyceraldehyde.Pathways are not closed, compounds come in and leave as well.Fatty acids are toxic and can not be stored. Hence, converted to triglycerols and fatty acid cleaves off glycerol to form 2C.Glycerol backbone is 3C. Glycerol is only part used in gluconeogenesis.Phospholipids ---- glycerol 3 phosphate which is oxidized to dihydroxy acetone phosphate.In fat synthesis, glycolysis is used to create fat as phospholipids are made vis glycolysis.Oxidation of glyceraldehyde creates H.Phosphate is attached to form ATP by phosphoglycerol kinase.3 phosphoglycerol -- 2 phosphoglycerol is enabled by phosphoglyceromutase which is regulated by acetylation. Acetylation inhibits this.Interaction between glycolysis and pentose phosphate pathway:PPP is an anabolic pathway and NADP in reduced form is