Dr. Saidunnisa, MD Professor of Biochemistry The Journey through the Hexose monophosphate Pathway!!!!

Learning objectives At the end of the session student shall be able to: The origin, differentiation and morphology of haemopoetic cells along with the metabolic pathways involved: Explain the steps of HMP (Hexose monophosphate shunt). Correlate the steps to defects in these pathways. (Lecture)

Case-1 A 25 year old African American was going on pleasure trip to Iraq . In preparation for his tour he is given a prophylactic dose of primaquine to prevent malaria. Several days after he began taking this drug he develops fatigue and hemolytic anemia. Why Discuss?

Case-2 Why?

Hexose Monophosphate shunt Other names: Pentose phosphate pathway Phosphogluconate oxidative pathway

It’s a shunt Glucose instead of going through glycolysis it is shunted through this pathway. 10% of glucose molecules per day are entering this pathway.

The pentose phosphate pathway has two main functions Production of NADPH Ribose -5-P Site : In the cytoplasm of specialized cells.

Generation of NADPH (Oxidative phase of HMP) mainly used for reductive syntheses of fatty acids. Not active in non-lactating mammary gland and has low activity in skeletal muscle.

Ribose-5-Phosphate (Non-Oxidative phase of HMP) Are required for the synthesis of the nucleotides and nucleic acids (DNA, RNA).

Reactions of the pentose phosphate pathway occur in the cytosol in two phases Phase-I : Oxidative, non-reversible phase Phase-II: Non-oxidative, reversible phase NADP+, not NAD +, is used as hydrogen acceptor

Don’t panic, you need not know all the reactions in detail; stay tuned!!!! Glucose-6-phosphate Dehydrogenase catalyzes oxidation of the aldehyde at C1 of glucose-6-phosphate, to a carboxylic acid in ester linkage (lactone). NADP+ serves as electron acceptor.

Non oxidative phase C5 + C5 C3 + C7 (Transketolase) C3 + C7 C6 +C4 (Transaldolase) C5 + C4 C6 + C3

Mechanism of reaction need not know Transfer of a 3-C

Regulation of pentose phosphate pathway Glucose 6-phosphate dehydrogenase is the rate limiting enzyme. This enzyme is regulated by availability of the substrate NADP+. Is stimulated by insulin.

Glutathione and NADPH What is glutathione? Why is it important? How is it related to NADPH?

Glutathione

Special functions of NADPH in RBC Reduced glutathione (GSH) is essential to maintain the integrity of RBC membrane. Is necessary to keep the iron in ferrous form i.e reduced (Fe+2) which can carry oxygen.

Pentose phosphate pathway protects cells against reactive oxygen species (ROS) Reduction of molecular O2 in a series of one-electron steps yields superoxide, hydrogen peroxide, hydroxyl radical, and water. The intermediate, activated forms of oxygen are known as reactive oxygen species (ROS)

Role of NADPH and glutathione in protecting cells against ROS Reduced glutathione (GSH) protects the cell by destroying hydrogen peroxide and hydroxyl free radicals by glutathione peroxidase . Regeneration of GSH from oxidized form (GS-SG) requires the NADPH (produced in the glucose 6- phosphate dehydrogenase reaction) by glutathione reductase .

So, what happens if glucose 6-phosphate dehydrogenase is defective? Insufficient production of NADPH. Which translates into insufficient glutathione. Is this a medical problem? YES

G6PD Deficiency Is the most common human enzyme deficiency in the world. Over 400 million people worldwide affected with highest prevalence in individuals of African, Mediterranean and Asian heritage. An X-linked disorder that therefore typically affects men.

G6PD Deficiency Most G6PD-deficient individuals are asymptomatic. But when certain drugs are taken such as (sulfa antibiotics, antimalarials like primaquine) and toxic ingredient of fava beans, stimulate peroxide formation leading to sudden damage to RBC and clinical manifestations occur.

Precipitating factors in G6PD deficiency.

Glucose-6-phosphate dehydrogenase deficiency causes hemolytic anemia Mutations in glucose 6-phosphate dehydrogenase, results in impairment of NADPH production. Detoxification of H2O2 is inhibited, and cellular damage results - leads to erythrocyte membrane breakdown and hemolytic anemia and jaundice.

Drug induced hemolytic anemia

Grown worldwide Important in Middle East High in protein Fava beans rich in two glycosidic compounds: Vicine, Convicine Upon ingestion, these are converted into divicine and isouramil These two compounds deplete reduced glutathione leading to formation of free radicals and H202 that cause hemolysis.

Glucose-6-phosphate dehydrogenase deficiency and resistance to malaria G-6PD frequently in Africans-protects them from malaria. Plasmodium falciparum is dependent on HMP shunt and reduced glutathione for their optimum growth in RBC. One theory to explain this, is that cells infected with the Plasmodium parasite are cleared more rapidly by the spleen.

Glucose-6-phosphate dehydrogenase deficiency causes hemolytic jaundice Normal value of G6PD in RBC is 6-12u/g of Hb.

Learning checks!!

Learning check!!

Brain storming!!! What is the rate limiting enzyme for the oxidative (irreversible) step? And what are its products? What is the enzyme for the non-oxidative step and its important product? Glucose-6-P dehydrogenase Transketolase

What will Glucose-6-phosphate dehydrogenase deficiency cause?

G6PD deficiency What type of genetic disease? What type of anemia and precipitating factors? Are their clinical manifestations in most individuals with a deficiency mutation?

Wernicke-Korsakoff syndrome Wernicke first described an illness consisted of ataxia, paralysis of eye movements (nystagmus) and mental confusion. SS Korsakoff Russian psychiatrist described disturbance of memory in alcoholism. Genetic disorder associated with transketolase activity results in Wernicke-Korsakoff syndrome. These manifest in alcoholics whose diets are deficient in vitamin thiamine. Transketolase is measured in RBC is an index of thiamine status of an individual

Recommended reading: Basic medical Biochemistry –A clinical approach – 3rd edition. Text Book of biochemistry for medical students DM Vasudevan. Lippincott's text book of Biochemistry. Google images