BMS100_BCH2.02-F22_CHO metabolism_STUDENT.pptx

BCH 2.02 Carbohydrates metabolism In-class Dr. Rhea Hurnik BMS100 Plan Pre-learning Review Glycolysis and Gluconeogenesis In-class Glycolysis continued Regulation of glycolytic Metabolism of other monosaccharides E.g., fructose Gluconeogenesis continued Regulation of gluconeogenic enzyme Pent...

BCH 2.02 Carbohydrates metabolism In-class Dr. Rhea Hurnik BMS100 Plan Pre-learning Review Glycolysis and Gluconeogenesis In-class Glycolysis continued Regulation of glycolytic Metabolism of other monosaccharides E.g., fructose Gluconeogenesis continued Regulation of gluconeogenic enzyme Pentose phosphate shunt General pathway Regulation Glycogenesis & Glycogenolysis General pathways Regulation Learning outcomes •Diagram the connections between glycolysis, gluconeogenesis, pentose phosphate shunt, glycogenesis, and glycogenolysis •Glycolysis Briefly review glycolysis, including reactants & products, enzyme names, and any allosteric and hormonal regulators of the enzymes. •Irreversible steps of glycolysis are testable on the next quiz, test, and final exam, include substrate, product, enzyme, and enzyme regulation Describe the metabolism of fructose and how it feeds into glycolysis •Gluconeogenesis Briefly review gluconeogenesis, including reactants & products, enzyme names, and any allosteric and hormonal regulators of the enzymes •Bypass reactions of gluconeogenesis are testable on the next quiz, test, including substrate, product, enzyme, and enzyme regulation Note: Gluconeogenic substrates are testable on the final exam. •Describe the pentose phosphate shunt include starting substrates and glycolytic products. Describe the key product of the pentose phosphate shunt, including NADPH and pentose phosphates Describe the allosteric and hormonal regulation of glucose-6-Phosphate dehydrogenase •Outline glycogenesis and glycogenolysis pathways, including key enzymes involved. Describe allosteric and hormonal regulation of glycogen synthase and glycogen phosphorylase Cytoso l Glucos e AT P *NAD H Glycoge n AT P Glucose 6P Pyruva te Oxaloacetat e AT P *NAD H Pyruvat e Oxaloacetat e CO Mitochondri 2 a *NADH, FADH2 O2 *NADH, FADH2 NADP H Pentose -PSugars Triglyceride s Glycer ol Fatty AcylCoA Fatty Acids *NAD H AcetylC oA *NAD H, FADH2 Citra te Citric Acid Cycle AT P Electron Transport Chain Keton es Fatty AcylCoA AT P H2O AT P AT P Acety lCoA Metabolism Review  Once Glucose is converted to Glucose-6-P (trapped molecule) it can enter several pathways?  Glycolysis  Which enzyme is needed to commit glucose-6P to glycolysis?  PFK1 (key controlling enzyme)  Pentose Phosphate Shunt (irreversible G6P--> pentose)  Glycogenesis Glycolysis: Regulation • The enzymes of the three irreversible reactions are also the main regulated enzymes of glycolysis  What are they?  PFK1 (fructose 1 phosphate  fructose 1,6 bisphosphate)  Hexokinase/glucokinase (glucose  G6P)  Pyruvate kinase (PEP  Pyruvate) Glycolysis: Allosteric Regulation - Hexokinase • What reaction does Hexokinase catalyze?  Hexokinase in muscle; sensitive to cell’s need for glucose and will thus be inhibited when there is a lot of glucose-6-phosphate product. • Reversibly regulated by glucose-6-phosphate • This is an example of product inhibition  When cellular concentrations of glucose-6-P rise above normal, Hexokinase is temporarily inhibited to bring rate of glucose-6-P into balance with its rate of utilization Glycolysis: Allosteric Regulation – PFK1  What reaction does PFK-1 catalyze  PFK-1 is under complex allosteric regulation  1. ATP inhibits PFK-1 by binding to an allosteric site  Lowers the affinity of the enzyme for fructose-6-P  AMP and ADP relieve the inhibition of PFK-1  Explain why this makes sense logically  2. Citrate  Key intermediate in CAC  High citrate levels inhibit PFK-1  High citrate serves a signal that the cell is meeting its current needs for energy  Citrate leaves mitochondrial matrix to be converted to fat if glucose is very high; it being sensed in cytosol is a key regulator for PFK to inhibit/downregulate glycolysis (too much CHO; excess converted to fats) Glycolysis: Allosteric Regulation – PFK1  PFK-1 is under complex allosteric regulation  3. Fructose 2,6-bisphosphate  Most significant allosteric regulator  Activates PFK-1  Fructose-2,6-bisphosphate (between fructose 6 phosphate and fructose 1,6 bisphosphate) enhances the affinity of PFK-1 for fructose-6-P  Stimulates glycolysis  Inhibits FBP-1, slowing gluconeogenesis  Fructose bisphosphatase (enzyme needed to convert fructose 1,6 bisphosphate back to fructose 6 phosphate in gluconeogenesis) Glycolysis: Allosteric Regulation – PFK1  PFK-1 is under complex allosteric regulation  3. Fructose 2,6-bisphosphate continued  Fructose 2,6-bisphosphate is formed by phosphorylating Fructose-6-P, catalyzed by phosphofructokinase-2 (PFK-2) CHOrich meal Blood glucose rises Controlling enzymes • Addition of phosphate shuts down enzyme • Removal of phosphate turns on enzyme • During FED state, phosphates are removed from enzymes • During FASTED state, phosphates are added to turn on enzyme  Insulin supports phosphate removal of enzymes (fed); glycolysis supported  Glucagon supports phosphate covalent modification on enzymes (fasted); glycolysis not supported (want to store glucose in this case) Glycolysis: Allosteric Regulation – Pyruvate Kinase  What reaction does pyruvate kinase catalyze?  Pyruvate kinase is allosterically regulated by:  Inhibition: try to explain each logically  1. ATP  2. Acetyl-CoA  3. Fatty acids  Activation:  Fructose 1,6-bisphosphate Glycolysis: Hormonal Regulation  Glycolysis is regulated hormonally by insulin and glucagon.  Insulin promotes the transcription of glucokinase, phosphofructokinase-1, and pyruvate kinase  Reminder: Insulin also promotes the activation of PFK-1 ( & inhibition FBP-2)  Covered on slide 21  Glucagon reduces the expression of hexokinase, PFK1, and pyruvate kinase  Glucagon promotes the activation of FBP-2 (& inhibition PFK-2) Glycolysis Review Glycolytic enzyme Activated by Inhibited by Glucokinase Insulin Glucagon Hexokinase Insulin Glucagon High levels of G6P PFK-1 Fructose 6 phosphate  fructose 1,6 bisphosphate F-1,6-BP being made * most important one Citrate present in cytosol and ATP Pyruvate Kinase PEP  pyruvate Insulin Glucagon ATP, acetyl CoA and FA Metabolism of other important sugars • Fructose, Mannose, and galactose are converted into glycolytic intermediates  We are only going to look at Fructose metabolism in more detail • Predominately occurs in the liver Fructose Fructose -1Phosphate DHAP + glyceraldehyde Glycolysi s Lipogenesis Fructose metabolism Fructose Fructose metabolism – thinking questions • Which monosaccharide is metabolized faster, glucose or fructose, why?  Fructose • Can be more readily made into fat Gluconeogenesis Regulation • The first two bypass reactions are the main regulated reactions of gluconeogenesis  What were these bypass reactions? • G6P  glucose • Pyruvate  oxaloacetate  Oxaloacetate  PEP  PEP  pyruvate, eventually glucose Gluconeogenesis: Allosteric Regulation – Pyruvate carboxylase  Pyruvate can be converted either to:  A) acetyl CoA  enter into CAC  B) Oxaloacetate  enter into GNG  What enzyme catalyzes conversion of pyruvate to oxaloacetate  C) Lactate (Cori cycle)  Acetyl CoA is an allosteric activator of pyruvate carboxylase Gluconeogenesis: Allosteric Regulation – FBP-1  What reaction does FBP-1 catalyze  Allosterically inhibited by AMP  Allosterically inhibited by Fructose 2,6bisphosphate Blood glucose is low Gluconeogenesis: Hormonal Regulation  Gluconeogenesis is regulated hormonally by insulin and glucagon.  Glucagon promotes activation of FBP-2, lowering levels of Fructose-2,6bisP  Previous slide  Glucagon can also induce the transcription of PEP carboxykinase  Fasting state  making PEP to eventually form glucose Gluconeogenesis Review Gluconeogenesis enzyme Pyruvate carboxylase PEP carboxykinase FBP-1 Activated by Inhibited by Metabolism Review  Once Glucose is converted to Glucose-6-P it can enter several pathways?  Glycolysis  Which enzyme is needed to commit glucose-6P to glycolysis?  Pentose Phosphate Shunt  Glycogenesis Pentose Phosphate Shunt • One fate of Glucose-6-phospate is the pentose phosphate Shunt • Alternative metabolic pathway for glucose that “shunts” molecules into and out of glycolysis • Purpose: • Generates two main products:  NADPH  Ribose-5-Phosphate (“pentose phosphates) • Also generates Fructose-6-Phosphate and glyceraldehyde-3-Phosphate, which can feed back into glycolysis  Why do you think it is called a “shunt?”  Location? Pentose Phosphate Shunt Glucose-6-P 6-Phosphoglucono-𝛿 lactone 6Phosphogluconat e • Two phases:  Oxidative • Irreversibl e reactions Ribose 5P Glyceraldehyd e-3-P D-Ribulose 5phosphate Sedoheptuloas e-7-P  Nonoxidative Xylulose-5-P • Reversible reactions Glyceraldehyde-3-P Erythrose-4-P Fructose-6-P Fructose-6-P Oxidative phase • Glucose-6-phosphate  Ribulose-5Phosphate  Generates 2 NADPH • Functions:  Fatty acid synthesis – more to come next week  Reduces glutathione (antioxidant)+ CO 2 3 Glucose-6-P 6-Phosphoglucono-𝛿 lactone 6Phosphogluconat e D-Ribulose 5phosphate NADPH: antioxidant • Glutathione is an antioxidant  3 amino acid peptide • Glycine-cystineglutamate  Neutralizes (reduces) hydrogen peroxide to water by donating H’s  NADPH regenerates glutathione by replacing the donated H’s Oxidative phase regulation • Glucose-6-phosphate  Ribulose-5Phosphate  Rate-limiting step: • Glucose-6-P  6-phosphoglucono-𝛿-lactone  Enzyme: glucose-6-Phosphate dehydrogenase (G6PD) • Regulated by ratio of NADPH:NADP+ • High NADPH:NADP+ ratio inhibits G6PD • Also inhibited by high levels of Acyl CoAs – why? • Upregulated by insulin 3 Glucose-6-P 6-Phosphoglucono-𝛿 lactone 6Phosphogluconat e D-Ribulose 5phosphate G6PD deficiency • G6PD deficiency is an X-linked trait • Therefore, is it more common in men or women?  Men  FYI - Highest prevalence in those of Middle Eastern, Mediterranean, and tropical African or Asian descent • Protect against certain kinds of malaria • Results in hemolytic anemia when an individual is exposed to oxidant stress  Ie. Some drugs, infections, fava beans Reactions all FYI Non-oxidative phase • Riboluse-5-P is converted to Ribose-5-P  What do you suppose Ribose-5-P can be used for? Ribose 5P Glyceraldehyd e-3-P Sedoheptuloas e-7-P Xylulose-5-P • OR into glycolytic intermediates  Which two molecules are the glycolytic intermediates? D-Ribulose 5phosphate Glyceraldehyde-3-P Erythrose-4-P Fructose-6-P Fructose-6-P Metabolism Review  Once Glucose is converted to Glucose-6-P it can enter several pathways?  Glycolysis  Which enzyme is needed to commit glucose-6P to glycolysis?  Pentose Phosphate Shunt  Glycogenesis Glycogen metabolism • Glucose is stored in polymeric form as glycogen mostly in the liver and skeletal muscle. • High glucose/energy levels will triggers glycogen synthesis  Glycogenesis • Glucose can be rapidly delivered to the blood stream when needed by degradation of glycogen in the liver  Glycogenolysis Glycogenesis • Glucose is transferred onto a growing chain of glycogen as UDP-glucose  Steps: UTP Glucose-6-P Phosphoglucomutas e Enzyme names FYI UDP-glucose + UDP-glucosePPi Glucose-1-P pyrophosphoryla se Glycogenesis • UDP-glucose is added to an existing strand of glycogen by the enzyme glycogen Glycogennsynthase + UDP-glucose → Glycogen • Glucose is n+1 added to the nonreducing end in what type of link? + UDP Glycogenesis - branching • Branching enzyme catalyzes the transfer of 4-8 glucose residues to a branch point  Branching enzyme is also called amyloα(1,41,6) glucosyl transferase Glycogenesis Primer • Glycogen synthase cannot initiate a new glycogen chain de novo, it requires a primer  Primer contains pre-formed (alpha14) polyglucose chain with at least 4-8 glucose residues  Found within glycogenin  Contains the primer and the enzyme needed to build the primer Glycogenesis Primer creation - visual • Notes Glycogenolysis • Glycogenolysis is a catabolic progress  Breakdown of glycogen into glucose units  Occurs in the liver and muscle Glycogenolysis • Glucose is removed from nonreducing ends of glycogen  Enzyme: glycogen phosphorylase • Sequentially cleaves α (1 4) linkages from the non-reducing ends until 4 units away from a branch point  Glucose is released as glucose-1-P • Once all chains degraded to within 4 units of a branch point, the molecule is called a limit dextrin Glycogenolysis • A de-branching enzyme has two functions:  1) transfers the outer 3 glucose residues from the branch to another non-reducing end (leaving only 1 reside behind at the branch point) • Aka Oligo-α(1,4)-α(1,4)glucotransferase  2) Removes the final glucose residue in the alpha(16) linkage • Aka Amylo-α(1,6)glucosidase Glycogenolysis • Glucose-1-P is converted to glucose-6-P  Enzyme: phosphoglucomutase • This is the same enzyme we saw in the first step of glycogenesis • The liver can then convert glucose-6-P into glucose with the enzyme glucose-6phosphatase  Hint: This enzyme catalyzes the third bypass reaction of gluconeogenesis  Muscles do not have this enzyme, why? What does the muscle do with glucose-6-P instead? Glycogen metabolism regulation • Glycogen synthase and glycogen phosphorylase are the two regulated enzymes in glycogen metabolism  Allosteric regulation: • Glycogen synthase regulation  Allosterically activated by glucose-6-P • Glycogen phosphorylase regulation  Allosterically inhibited by: • Glucose-6-P • ATP • Free glucose (in the liver only)  Allosterically activated by AMP (muscle only) Glycogen metabolism regulation - Hormonal • Glycogen synthase and glycogen phosphorylase can also be regulated by covalent modification  Glycogen synthase is de-activated by phosphorylation  Glycogen phosphorylase is activated by phosphorylation • Phosphorylation is catalyzed initially by the same protein, protein kinase A (PKA)  How is PKA activated? • This prevents both pathways from running at the same time activated Review Inhibited • Glycogen metabolism Protein kinase A ATP Protein kinase A ADP Glycogen phosphoryl ase kinase Glycogen phosphoryl ase kinase ATP Glycogen phosphoryl ase ATP Glycogen synthase P ADP Glycogen synthase ADP Glycogen phosphoryl ase Glycogenol ysis P No glycogenesi s P activated Review Inhibited • Glycogen metabolism continued Protein kinase A ATP Protein kinase A ADP Glycogen phosphoryl ase kinase Glycogen phosphoryl ase kinase ATP Glycogen phosphoryl ase ATP Glycogen synthase P ADP Glycogen synthase ADP Glycogen phosphoryl ase Glycogenol ysis P No glycogenesi s P Glycogen metabolism Regulation - Hormonal • Covalent modification of glycogen metabolism is under hormonal control  In the presence of glucagon (and epinephrine): • Glucagon binds to its GCPR • G⍺s activates adenylyl cyclase  cAMP levels rise • PKA phosphorylates glycogen synthase, rendering it INACTIVE  Glycogenesis is inhibited Glycogen metabolism Regulation Hormonal • Covalent modification of glycogen metabolism is under hormonal control In the presence of glucagon (and epinephrine): • PKA phosphorylates glycogen phosphorylase kinase, rendering it ACTIVE • Glycogen phosphorylase kinase phosphorylates glycogen phosphorylase, rendering it ACTIVE • Glycogen metabolism Regulation - Hormonal • Covalent modification of glycogen metabolism is under hormonal control  In the presence of Insulin • Insulin promotes the breakdown of cAMP and thus inactivation of PKA • Insulin activates protein phosphatase 1, which removes the phosphate group from glycogen synthase, rendering it active • Glycogenesis is promoted Glycogen metabolism - Hormonal •Regulation Covalent modification of glycogen metabolism is under hormonal control In the presence of insulin • Insulin promotes the breakdown of cAMP and thus inactivation of PKA • Insulin activates protein phosphatase 1, which removes the phosphate group from: • Glycogen phosphorylase kinase & Glycogen Glycogenolysis is inhibited References • Alberts et al. Molecular Biology of the Cell. Garland Science. • Lehninger Principles of Biochemistry 4th ed. Figure 21-36. Page 818 • Images:  https://upload.wikimedia.org/wikipedia/ commons/b/bf/ Pentose_phosphate_pathway_Kor.png Study Questions • Draw out a basic schematic linking glycolysis, gluconeogenesis pentose phosphate shunt, and glycogenesis, glycogenolysis  In your schematic, include the key regulated reactions and enzymes for each pathway • Add the allosteric regulators to your diagram  How does Fructose to feed into glycolysis