MHS_PENTOSE PHOSPHATE PATHWAY_Nayyar.pdf

Pentose Phosphate Pathway Tultul Nayyar, Ph.D., MSCI Associate Professor and Director Master of Health Sciences Program West Basic Science, Room 3210 Phone: 615-327-6898 (W) 615.944.0824 (C) email: [email protected] The pentose phosphate pathway is a shunt • The pathway begins with the glycolytic i...

Pentose Phosphate Pathway Tultul Nayyar, Ph.D., MSCI Associate Professor and Director Master of Health Sciences Program West Basic Science, Room 3210 Phone: 615-327-6898 (W) 615.944.0824 (C) email: [email protected] The pentose phosphate pathway is a shunt • The pathway begins with the glycolytic intermediate glucose 6-P • It reconnects with glycolysis because two of the end products of the pentose pathway are glyceraldehyde 3-P and fructose 6-P (which are the members of glycolytic pathway) • The pentose pathway is often therefore, referred to as a Hexose monophosphate shunt (viewed as a shunt of glycolysis) It’s a shunt and takes place in the cytosol What does the pentose phosphate pathway achieve? Yields: NADPH (to be used in anabolic reactions requiring electrons) • Fatty acids and steroids synthesis • Reduced glutathione synthesis (for antioxidant activity to protect cells from oxidative damage) Ribose (pentose sugar) • Nucleotide synthesis • DNA • RNA * Tissues with active pentose phosphate pathway Pentose Phosphate Pathway Divided into two phases 1. Oxidative Phase: Irreversible reactions Generates NADPH 2. Nonoxidative Phase: Reversible reactions Generates ribose precursors * Pentose phosphate pathway Reactions Oxidative Phase: Regulatory enzyme 5 carbon atoms Oxidative Phase (reactions 1-3) * Reaction 1: Glucose-6-P dehydrogenase Regulatory enzyme  Glucose-6-P dehydrogenase enzyme is highly specific for NADP+  First site for NADPH formation Reaction 2: * Reaction 3:  Second site for NADPH formation Nonoxidative Phase: Non-0xidative reactions (reactions 4-8) (~Reversible reactions) Ribulose -5-P (keto-pentose sugar) converts to: Xylulose-5-P (keto-pentose sugar) Reaction 4: by ribulose-5-P-epimerase OR Ribulose -5-P (keto-pentose sugar) converts to: Ribose-5-P(aldo-pentose sugar) Reaction 5: by ribulose-5-P-isomerase CH2OH Ribulose-5-PO4 C=O H-C-OH H-C-OH CH2OP Ribulose-5-PO4 Epimerase CH2OH C=O HO-C-H H-C-OH CH2OP Xylulose-5-PO4 Ribulose-5-PO4 Isomerase CHO H-C-OH H-C-OH H-C-OH CH2OP Ribose-5-PO4 Reaction 6: Xylulose-5-P (C5)+ Ribose-5-P (C5) forms Sedoheptulose 7-P (keto-heptose sugar, C7) + Glyceraldehyde-3-P (ald0-triose, C3) by transketolase enzyme (by breaking the bond between C2C3 of keto sugar,i.e. transfers two C unit, C1C2)  Requires thiamin pyro phosphate (TPP, Vitamin B1) as coenzyme Remember! Thiamine pyrophosphate (TPP – a coenzyme derived from thiamine/B1) requiring enzymes: • Transketolase (catalyzes the conversion of pentose phosphate pathway sugars into glyceraldehyde 3phosphate – which is a glycolysis intermediate) • Pyruvate dehydrogenase (PDH - catalyzes the oxidation & decarboxylation of pyruvate to acetyl CoA) • Alpha ketoglutarate dehydrogenase (catalyzes the oxidation of alpha ketoglutarate into succinyl CoA in the Krebs cycle) Thiamine deficiency affects severely brain functions : Manifestations: Confabulation (production of fabricated, distorted or misinterpreted memories without the intention to deceive), psychosis, ataxia (uncoordinated movement), nystagmus (rapid eye movement) & ophthalmoplegia (eye muscle paralysis). * Among the coenzymes needed for pyruvate dehydrogenase is thiamine pyrophosphate, also known as vitamin B1. Lack of this vitamin leads to beri-beri, a debilitating disease often suffered by prisoners of war. Which other enzyme complex in the citric acid cycle also has a requirement for thiamine pyrophosphate? A. Citrate synthase B. Citrate isomerase C. Isocitrate oxidoreductase D. α-Ketoglutarate oxidoreductase E. Malate oxidoreductase Reaction 7: Sedoheptulose 7-P + Glyceraldehyde-3-P forms Fructose-6-P (keto-hexose sugar, C6) + Erythrose-4-P (aldo-tetrose dugar, C4) by transaldolase enzyme (by breaking the bond between C3-C4 of keto sugar (i.e. transfers three C unit, C1C2C3) Reaction 8: Xylulose-5-P (keto-C5) + Erythrose-4-P (aldo-C4) forms Fructose-6-P (keto-hexose sugar, C6) + Glyceraldehyde-3-P (ald0-triose, C3) by transketolase enzyme (by breaking the bond between C2-C3 of keto sugar, i.e., transfers two C unit, C1C2)  Requires thiamin pyro phosphate (TPP, Vitamin B1) as coenzyme  Transketolase and transaldolase enzymes actions make the link to glycolysis  Glyceraldehyde 3-phosphate  Fructose 6-phosphate * Pentose Phosphate Pathway can account for complete oxidation of glucose *  Because • 3 molecules of glucose-6-P (i.e., 3 x 6 = 18C) to make 3 molecules of CO2 (3C)+ 3 molecules of 5C-sugars (15C) 2 molecules of glucose-6-P (12C)+ 1 mol glyceraldehyde 3-P (3C) 6 G6P + 12 NADP+ + 6 H2O → 5 G6P + 12 NADPH + 12 H+ + 6 CO2 OR G6P + 12 NADP+ + 6 H2O → 12 NADPH + 12 H+ + 6 CO2 Regulation of the Pentose Phosphate Pathway * • Glucose 6-phosphate dehydrogenase is the regulatory enzyme • NADPH is a potent competitive inhibitor of the Glucose 6phosphate dehydrogenase enzyme • Usually the ratio NADPH/NADP+ is high so the enzyme is inhibited • But, with increased demand for NADPH, the ratio decreases, and enzyme activity is stimulated • Since the concentrations of the non-oxidative portion of the pentose pathway are reversible, the products and reactants can shift depending on the metabolic needs of a particular cell or tissue Glutathione - NADPH link to protect cell * • Glutathione is a tri-peptide composed of glutamate, cysteine, and glycine • The oxidized glutathione (G-S-S-G) is converted to reduced glutathione (G-SH) by glutathione reductase in presence of NADPH (which comes from pentose pathway) • The GSH is used to reduce peroxides, (Reactive Oxygen Species, ROS) formed during partial reduction of molecular oxygen in the ETC _ _ _ • O2 → O2 → O2 2 2O2 + 2H+ → O 2 + H2O2 • ROS damage macromolecules (DNA, RNA, and protein) and ultimately lead to cell death Glutathione and Erythrocytes * • GSH is extremely important particularly in the highly oxidizing environment of the red blood cell • RBCs have no mitochondria and are totally dependent on NADPH from the pentose phosphate pathway to regenerate GSH from GSSG by the enzyme glutathione reductase • GSH maintains cysteine residues in hemoglobin and other proteins in a reduced state • GSH is essential for normal RBC structure and keeping hemoglobin in Fe++ state • Individuals with reduced levels of GSH are subject to hemolysis • This is often clinically seen as black urine under certain conditions * Conditions for hemolytic anemia related glucose-6-p dehydrogenase (G6PD) deficiency • The ingestion of oxidative agents that generate peroxides or reactive oxygen species (ROS) • Individuals with G6PD deficiency can not produce sufficient GSH to cope with the ROS (because synthesis of NADPH gets affected) • Proteins become cross linked leading to Heinz body formation and cell lysis • G6PD deficiency is an inheritable X-linked recessive disorder • Approximately 10-14% of the male African American population is affected G6PD Deficiency leads to NADPH deficiency *** G6PD deficient individual : • when exposed to anti-malarial drugs (Primaquine): results in increased cellular production of superoxide and hydrogen peroxide (aka Primaquine sensitivity) • Hemolytic anemia • increased susceptibility to bacterial infections (phagocytes use NADPH for NADPH oxidase to make superoxide anions used to kill pathogens) • sickle cell anemia • when exposed to Other chemicals also known to increase oxidant stress • Sulfonamides (antibiotic) • Asprin and NSAIDs • Quinadine and quinine • Napthlane (mothballs) • Fava beans (vicine & isouramil) • G6PD deficiency is an X-linked recessive illness characterized by episodic *** hemolytic anemia that is most frequently induced by oxidative stress from infections or drugs. Which enzyme is most directly impaired in the Individual suffering from this illness? Catalase, Glucokinase, Glutathione peroxidase, or superoxide dismutase? Individuals with G6PD deficiency are less capable of generating the NADPH required to reduce glutathione. In turn, the lack of reduced glutathione reduces the ability of glutathione peroxidase to break down hydrogen peroxide, a reactive oxygen species that accumulates during infections and certain drug treatments. The accumulation of hydrogen peroxide leads to hemoglobin denaturation and membrane damage in red blood cells, resulting in hemolytic anemia, with the formation of Heinz bodies. Both catalase and superoxide dismutase also remove reactive oxygen species but are not directly affected by G6PD deficiency. • A man is given antibiotics to treat a bacterial infection and develops an episode of red blood cell lysis. Further studies show weakness of the plasma membrane and Heinz bodies (collections of oxidized hemoglobin). Which of the following enzymes is most likely defective in this patient? Fructose-1,6-bisphosphatase, Glucose-6-phosphate dehydrogenase, Hexokinase or Pyruvate kinase * Glucose catabolism: Glycolysis vs. Pentose pathway Glycolytic pathway Pentose pathway Utilizes NAD Not produces CO2 Generates ATP Utilizes NADP Produces CO2 Not generates ATP