PENTOSE PATHWAY&

ANTIOXIDANTS

BIOC 460 - DR. TISCHLER LECTURE 26

OBJECTIVES

1. For the pentose phosphate pathway:a. describe the oxidative and non-oxidative branches b. describe how the oxidative branch is regulatedc. distinguish between the 3 modes in terms of the roles of the

potential endproducts of each mode.

2. Describe the consequences of thiamine deficiency

3. In relation to antioxidant function in the body:a. list the major active (reactive) oxygen species, identify the

antioxidant which reduces that species.b. describe the metabolism of glutathionec. identify the enzymes that remove peroxides and superoxide

radicals from a cell and name their cofactor.d. describe the relationships between the components of the

antioxidant cascade including the reactions involved.e. discuss why a defect of glucose-6-phosphate dehydrogenase in the red blood cell might lead to loss of membrane integrity.

PHYSIOLOGICAL PREMISE

Do you have a partial enzyme deficiency about which you are unaware? There are circumstances where an individual may have such a partial deficiency but be unaware of the fact until a physiological event shifts the balance of metabolic processes. For example, individuals with malaria are given a drug called primaquine. When the body metabolizes primaquine it increases the demand for production of NADPH in most cells. A major source of NADPH is the glucose-6-phosphate dehydrogenase (G6PDH) reaction in the pentose phosphate pathway. In the red blood cell, this pathway is essential for removing peroxides, which can oxidize lipids in the plasma membrane causing the cell to become more fragile. Stressing the system with primaquine in an individual with a partial deficiency of G6PDH will lead to red cell destruction and hence the individual becomes anemic.

Functions of Pentose Phosphate Pathway

1) NADPH for biosynthetic pathways (e.g., synthesis of fatty acids and cholesterol);

2) NADPH for maintaining glutathione in its reduced state (see discussion of glutathione later);

3) Pentose sugar for synthesis of nucleic acids

glycolytic intermediates

Glucose 6-P 6-PhosphogluconateNADP NADPH

Glucose-6-P-DH

Xylulose 5-P Ribose 5-P

OxidativeBranch

Non-oxidativeBranch

Nucleicacids

Sedoheptulose-7-P

Erythrose 4-P

Transketolase

Transaldolase

Glyceraldehyde 3-P

Fructose 6-PFructose 6-P

Glyceraldehyde 3-P

TPPTransketolase

Ribulose 5-P CO2

NADPH

NADP6-Pgluconate DH

Figure 1. The pentose phosphate pathway containing an oxidative and a non-oxidative branch

Ribulose 5-P

Xylulose 5-PRibose 5-P

Sedoheptulose 7-P

Erythrose 4-P

Transketolase

Transaldolase

Glyceraldehyde 3-P

Fructose 6-PFructose 6-P

Glyceraldehyde 3-PT

ran

sk

eto

las

e

Figure 2. Using the non-oxidative branch of the pentose pathway to produce ribose-5-phosphate for the nucleic acid pathways (Mode 1).

Ribose-5-P is the sugar required for the synthesis of nucleic acids

Nucleic acids

Non-oxidativeBranch

Glucose 6-P

Ribulose 5-P

6-Phosphogluconate

Ribose 5-P

NADP NADPH

CO2 NADPH

NADP

Figure 3. Using the oxidative branch of the pentose pathway to produce NADPH for biosynthetic reactions and ribose-5-phosphate for producing nucleic acids (Mode 2).

Nucleicacids

OxidativeBranch

OxidativeBranch

Non-oxidativeBranch

back to glucose-6-P or to glycolysis

Glucose 6-P (3)

Ribulose 5-P (3)

6-Phosphogluconate

Xylulose 5-P (2) Ribose 5-P (1)

Sedoheptulose 7-P (1)

Erythrose 4-P (1)

NADP NADPH

CO2

NADPH

NADP

Glyceraldehyde 3-P (1)

Fructose 6-P (1)

Fructose 6-P (1)

Glyceraldehyde 3-P (1)

back to glucose-6-P or to glycolysis

Figure 4. Using the oxidative branch to produce NADPH for biosynthesis and returning ribulose-5-P to glycolytic intermediates (mode 3)

used by transketolase, PDH, KgDH

deficiency affects nucleic acid synthesis/energy metabolism

Wernicke-Korsakoff syndrome – observed in alcoholics due to poor diet

thiamine deficiency in individuals on high CHO diet (e.g., rice) causes beriberi

• patients tire easily• cardiac decompensation• energy depletion on high CHO diet

NUTRITIONAL PREMISE: THIAMINE (VITAMIN B1)

Brain atrophy due to Wernicke’s encephalopathy

Slide to be shown in class

Table 1. Reactive Oxygen Species and Antioxidants that Reduce Them

Reactive Species Antioxidant

Singlet oxygen 1O2 Vitamin A, vitamin E

Superoxide radical (O2-) superoxide dismutase, vitamin C

Hydrogen peroxide (H2O2)

Catalase; glutathione peroxidase

Peroxyl radical (ROO) Vitamin C, vitamin E

Lipid peroxyl radical (LOO) Vitamin E

Hydroxyl radical (OH) Vitamin C

O2

UV lightheme FeCoQ

1O2

NADPHor CoQ

O2-

H2O2

H+

H+

HOO

Lipid(LH)

L

H2O

O2

LOO

OH

Fe2+

H2O, H+

Figure 5. Pathways for the formation of reactive oxygen species

Superoxide dismutase

Haber-Weiss reaction; Fenton reaction

Singlet oxygen

Superoxide radical anion

Peroxyl radical lipid radical

lipid peroxyl radical

H2O2 glutathioneperoxidase

2 H2O

2 GSH

GSSGglutathionereductase

NADPH + H+NADP+

pentose pathway

Figure 6. Reactions of glutathione reduction and oxidation

SUMMARY OF ANTI-OXIDANT ENZYMES

Glutathione peroxidase: 2 GSH + H2O2 GSSG + 2 H2O

Uses selenium as a cofactor

Catalase : 2 H2O2 H2O + O2

Superoxide dismutase: 2 O2- + 2H+ H2O2 + O2

Mitochondrial - Mn2+ cofactor

Cytoplasmic – Cu2+-Zn2+ cofactors; mutations associated with familial amyotrophic lateral sclerosis (FALS)

Lipid Peroxidase: removes LOOH

selenocysteine in glutathione peroxidase intake may be related to lower cancer mortality

• cancer patients have lower plasma Se levels• risk may be higher in those with low Se intake• AZCC study – reduced incidence of prostate, colon, lung cancers

toxicity (> 1 mg/day) results in hair loss, GI upset, nerve damage

NUTRITIONAL CORRELATE: SELENIUM

Vit EredVIT Eox

Vit CredVIT Cox

LOOHlipid peroxyl radical LOO

Glutathionered

(GSH)

NADP+

NADPH + H+

Glucose-6-P Ribulose-5-P

Pentose phosphate pathway (rxn 8)

+ROOH

rxn 2

Glutathioneox

(GSSG)

H2O2

2H2O

hydroxyl radical (OH) superoxide radical (O2

-)

reduced products

Figure 7. Antioxidant cascade Reduced forms/reductionOxidized forms/oxidation

rxn 9

rxn 7

rxn 1

rxn 6

rxn 5

rxn 4

Medical Scenario:

If the antioxidant protective system in the red blood cell becomes defective, hemolytic anemia occurs; that is red blood cells undergo hemolysis and their concentration in the blood decreases. Such is the case if glucose 6-phosphate dehydrogenase is defective in the pentose phosphate pathway. In individuals whose glucose 6-phosphate dehydrogenase is defective, there is insufficient NADPH produced in red blood cells to maintain the ratio of reduced glutathione to oxidized glutathione at its normal value of well over 100. Hence, peroxides destroy the red cell membrane because of the limited protective mechanism in these cells.