Antioxidants Flashcards
Oxidative Branch of PPP
Glucose-6-phosphate —> 6-Phosphogluconate —> Ribulose-5-phosphate (Ribose-5-phosphate)
•The key enzyme in this branch and the rate-limiting and committed step in the pentose pathway is glucose-6-phosphate dehydrogenase.
- This reaction produces the first of two NADPH molecules formed per glucose-6-phosphate.
- Glucose-6-phosphate dehydrogenase also is the only site of regulation in the pathway being sensitive to feedback inhibition by NADPH.
- The second molecule of NADPH produced in the oxidative branch is a product of 6- phosphogluconate dehydrogenase, which catalyzes the decarboxylation of the hexose to a pentose sugar.
- Although formally the oxidative branch ends with ribulose-5-phosphate, when the oxidative branch is operative without the non-oxidative branch the endproduct is ribose-5-phosphate to provide the sugar moiety of nucleotides.
Nonoxidative Branch of PPP
- The non-oxidative branch, unlike the oxidative branch, is reversible depending on the needs of the cell.
- Ribose-5-phosphate is a precursor for synthesis of nucleic acids.
- The non-oxidative branch includes a variety of sugar intermediates containing four, five or seven carbons.
- Transketolase, an enzyme of clinical note, catalyzes two of the three reactions of this branch.
-Transketolase requires thiamine diphosphate (TDP) as a cofactor (prosthetic group) as well as Mg2+ ions.
- When the non-oxidative branch begins with ribulose-5-phosphate, the endproducts are the glycolytic intermediates glyceraldehyde-3-phosphate and fructose-6- phosphate.
- The pathway can be reversed by starting with these intermediates of glycolysis and ending with the production of ribose-5-phosphate.
Cell needs NADPH but not ribose-5-P?
- Glucose-6-phosphate (G6P) is produced via hexokinase in most tissues but by glucokinase in liver and pancreatic beta-cells.
- The G6P proceeds through the oxidative branch to ribulose-5-phosphate.
- This pentose is then converted to fructose-6-phosphate and glyceraldehyde-3-phosphate.
- These intermediates can then be metabolized to pyruvate in the glycolytic pathway or returned to G6P via the gluconeogenic pathway.
- Under these conditions the pathway is ‘shunting’ around part of glycolysis, hence its alternate name, the hexose monophosphate shunt.
- To achieve balance of the reactions in the non-oxidative branch two-thirds of the ribulose-5-phosphate must be used to form xylulose-5-phosphate with the remaining one-third being converted to ribose-5-phosphate.
Cell needs both NADPH and ribose-5-P?
•To meet this requirement, the cell operates only the oxidative branch of the pentose pathway. Thus ribulose-5-P is all converted to ribose-5-P. The ribose-5-P is used for the synthesis of nucleic acids.
Cell needs ribose-5-P for nucleic acid synthesis but does not require NADPH?
- Fructose-6-phosphate and glyceraldehyde-3-phosphate are provided from glycolysis for the non-oxidative branch.
- The non-oxidative branch proceeds in the opposite direction.
- Thus ribose-5-phosphate is produced from these glycolytic intermediates.
- Under these conditions the oxidative pathway is shut down by NADPH feedback inhibition.
Thiamine (Vitamin B1)
- The cofactor form of thiamine, a water-soluble vitamin, is thiamine diphosphate.
- Thiamine diphosphate is not only a cofactor for the transketolase reaction of the pentose phosphate pathway, but also for the pyruvate and alpha-ketoglutarate dehydrogenase reactions.
- Thus, thiamine deficiency affects both ribose-5-phosphate production for synthesis of nucleic acids and energy metabolism, especially of carbohydrates.
Thiamine Deficiency
- Wernicke-Korsakoff Syndrome
- Beriberi
Thiamine Deficiency - Wernicke Korsakoff Syndrome
•Wernicke-Korsakoff syndrome is caused by thiamine deficiency, though at one time it was believed to be due to chronic alcohol intoxication.
-The syndrome is often observed in alcoholics because their nutritional status is very poor.
- Patients with Wernicke disease exhibit eye signs (i.e., ophthalmoplegia), ataxia and mental changes.
- Korsakoff psychosis is associated with memory loss.
- The irreversible brain damage that ensues in this syndrome may be a consequence of decreased synthesis of nucleic acids, which are in high demand in nervous tissue.
- If the patient is treated in time with thiamine, then brain damage can be prevented.
Thiamine Deficiency - - Beriberi
- When thiamine deficiency occurs in individuals on a relatively high carbohydrate diet (e.g., ricebased diet), beriberi develops.
- Beriberi in Singhalese means “weakness”.
- The disease is associated with tiring easily, heavy feeling of the limbs, and cardiac decompensation.
-These symptoms are all related to energy depletion, since individuals on a high carbo-hydrate diet depend on both pyruvate and alpha-ketoglutarate dehydrogenase for the aerobic production of ATP from glucose.
- Wet beriberi, unlike the dry form, is associated with edema.
- The disease can be fatal as a result of heart failure or general failure of the circulatory system.
- Food sources of thiamine are many. The best sources are organ meats, lean meats, eggs, green leafy vegetables, nuts and legumes.
- However, polishing of white rice removes the thiamine stores, thus accounting for the high incidence of beriberi in countries where polished or refined white rice is a main food source.
Reactive Oxygen Species
- Toxic ROS are generated during the normal course of cellular metabolism. These substances damage cells by peroxidation of membrane lipids, modifying proteins, and breakage of DNA strands.
- ROS are inhibited through the action of antioxidants. The major ROS, in order of increasing reactivity, include
- singlet oxygen
- superoxide radical
- hydrogen peroxide (most common)
- peroxyl radical
- lipid peroxyl radical
- hydroxyl radical
•Radicals are extremely reactive, especially against compounds that contain double bonds, such as unsaturated lipids (mostly found in membrane phospholipids) and nucleic acid bases. In a typical cell there are about 1000 attacks of free radicals on DNA per day. When damage does occur in membranes and proteins, the oxidized molecules are recycled. The role of antioxidants is to eliminate ROS before they can cause damage.
ROS - Singlet Oxygen
- Singlet oxygen is generated from molecular oxygen and is found mostly in skin.
- Singlet oxygen reacts with biological molecules and generates oxygen radicals (e.g., superoxide).
- It is a relatively minor contributor to oxidative damage in cells.
- Antioxidant: Vitamin A, Vitamin E
ROS - Superoxide Radical Anion
- Superoxide radical anion is simply referred to as “superoxide”.
- Biologically it is associated with the mitochondrial electron transport chain or NADPH oxidase in activated neutrophils, eosinophils and macrophages.
- It is sometimes a product when oxygen contacts reduced iron in hemoglobin.
- Antioxidant: Superoxide Dismutase, Vitamin C
ROS - Hydrogen Peroxide
- Hydrogen peroxide plays a central role in the formation of ROS and is the most common of these substances.
- Hydrogen peroxide is produced from superoxide via superoxide dismutase and as a byproduct of the conversion of superoxide to hydroxyl radical.
- Antioxidant: Catalase, Gutatione Peroxidase
ROS - Hydrogen Peroxyl Radical
- Hydrogen peroxyl radical has a reactivity similar to that of hydrogen peroxide.
- These radicals participate in many of the same reactions as hydroxyl radicals but are less reactive.
- Antioxidant: Vitamin C, Vitamin E
ROS - Lipid Peroxyl Radical
- Lipid peroxyl radical formation depends on action of hydroxyl radicals.
- One of the primary targets of hydroxyl radicals is reaction with membrane phospholipids and molecular oxygen to form lipid peroxyl radical.
- Lipid peroxyl radicals oxidize membrane lipids and in so doing regenerate more hydroxyl radical to perpetuate the sequence of events.
- Antioxidant: Vitamin E
Antioxidant Defense - Glutathione
•Glutathione peroxidase:
- Glutathione, a tripeptide (glu-cys-gly), exists predominantly in its reduced form (GSH) when the cysteine residue has a free SH group.
- The oxidized (GSSG) form consists of two tripeptides linked by a disulfide bond [-S-S-] between the cysteine residues.
- GSH is oxidized to GSSG via glutathione peroxidase at the expense of hydrogen peroxide.
- Thus reduced glutathione acts as a natural antioxidant.
•Glutathione reductase:
- Glutathione, which is oxidized in the peroxidase reaction, is regenerated to its reduced form via glutathione reductase.
- The reductase contains bound FAD to transfer electrons from NADPH to GSSG.
- NADPH is derived from the pentose pathway.
- Recall that niacin is the vitamin precursor of NADPH.
Antioxidant Cascade
- Glutathione: The interrelationships between various antioxidants can be depicted as a cascade. GSH protects the cell against damage from peroxides. GSH then is regenerated ultimately by NADPH produced in the pentose phosphate pathway.
- Vitamin C: Besides reducing H2O2, GSH plays an additional antioxidant role through its ability to regenerate reduced vitamin C (ascorbic acid). Reduced vitamin C protects cells against the oxidant effects of both hydroxyl radical and superoxide radical anion. Reduced vitamin C also restores vitamin E to its reduced state. Vitamin C is also important for maintaining iron in its reduced state in certain enzymes (i.e., prolyl and lysyl hydroxylase) of collagen synthesis.
- If sufficient amounts of vitamin C are lacking in the diet, symptoms of scurvy will occur within 45 to 80 days.
- Normal adults require 10 mg/day to prevent scurvy.
- The RDA of 60 mg/day for normal adults is the estimated amount that can be retained in the body. Intake beyond this leads to excretion in the urine and can increase the production of oxalates that contribute to the formation of kidney stones as calcium oxalate.
•Vitamin E: The antioxidant role of reduced vitamin E is primarily to rid the cell of lipid peroxyl radical, though it also acts on singlet oxygen and peroxyl free radical. This free radical scavenging role of vitamin E is its only known physiological function. Vitamin E, a fat-soluble vitamin, actually is comprised of a family of eight related compounds.
- The major constituent of the family is alpha-tocopherol. Because vitamin E is widely distributed in foods, a deficiency in humans is rare and is usually secondary to lipid malabsorption.
- Pregnant and lactating women may require vitamin E supplementation.
- A deficiency of vitamin E leads to increased peroxidation of membrane lipids, decreased mitochondrial activity, mutations of DNA, and damage to cell transport processes.
- The classic symptom of vitamin E deficiency is fragility of erythrocytes.
