- # About the Author
- # About the web site
- # Our second web site
- # Question of the day
- A New Book of Biochemistry
- Acid Base Balance
- Animations Links
- Biochemical Techniques
- Biochemistry Quiz
- Biological Oxidation
- Chemistry of Carbohydrates
- Chemistry of Lipids and Eicosanoids
- Chemistry of Nucleotides
- Chemistry of Proteins
- Diabetes Mellitus
- Diet and Nutrition
- Facebook Group Posts
- Haem Synthesis and Degradation
- Hemoglobin and Hemoglobinopathies
- Liver Function Tests
- Metabolism – Carbohydrates
- Metabolism – Lipids
- Metabolism – Nucleotides
- Metabolism – Proteins
- Metabolism of Alcohol
- Molecular Biology
- Past Papers
- Power Point Presentations
- Practical Biochemistry
- Abnormal Urine
- Blood Glucose Estimation
- Blood Urea and Urea Clearance Estimation
- Normal Laboratory Reference Values
- Normal Urine Analysis
- Power point presentations
- Protein Precipitation Reactions
- Reactions of Carbohydrates
- Serum Creatinine and Creatinine clearance estimation
- Serum Total Protein estimation
- Practice Questions
- Quick revisions
- Renal Function Tests
- Semester Paper
- Students’ corner
- Water and Electrolyte balance and Imbalance
Regulation and significance of HMP pathway (Lecture-3)
Glucose-6-phosphate Dehydrogenase is the committed step of the Pentose Phosphate Pathway. Following factors affect the activity of this enzyme and thus influence the rate of this pathway-
I) Availability of Substrate– This enzyme is regulated by availability of the substrate NADP+(Figure-1) As NADPH is utilized in reductive synthetic pathways, the increasing concentration of NADP+ stimulates the Pentose Phosphate Pathway, to replenish NADPH. The inhibitory effect of low levels of NADP+ is exacerbated by the fact that NADPH competes with NADP+ in binding to the enzyme. The marked effect of the NADP+ level on the rate of the oxidative phase ensures that NADPH generation is tightly coupled to its utilization in reductive biosyntheses. This explains the higher rate of activity of HMP pathway in tissues involved in reductive biosynthesis.
Figure-1- NADPH is the reduced form of NADP+(Nicotinamide Adenine Dinucleotide Phosphate)
2) Induction and repression– The synthesis of glucose 6-phosphate dehydrogenase and 6 -phosphogluconate dehydrogenase (Figure-2) may also be induced by insulin in the fed state, when lipogenesis increases. Excessive carbohydrate ingestion thereby leads to more NADPH generation. The synthesis of these enzymes is decreased in the fasting state.
The non oxidative phase of HMP pathway accomplishes conversion of the 5-C ribulose-5-phosphate to the 5-C product ribose-5-phosphate, or to the 3-C glyceraldehyde-3-phosphate and the 6-C fructose-6-phosphate. This phase is regulated by the flow of substrates.
Figure-2 Glucose-6-P dehydrogenase and 6- Phosphogluconate dehydrogenase are NADP+ specific enzymes . In tissues of high NADPH use, the enzyme activities are stimulated, as the inhibition by NADPH is overcome, with the resultant overall activation of the pathway.
Significance of HMP pathway
a) Biological Significance
The pentose phosphate pathway is primarily an anabolic pathway that utilizes 6 carbons of glucose to generate 5 carbon sugars and reducing equivalents. However, this pathway does oxidize glucose and under certain conditions can completely oxidize glucose to CO2 and water. The primary functions of this pathway are:
1. To generate reducing equivalents, in the form of NADPH, for reductive biosynthesis reactions within cells. The examples of reactions requiring NADPH (Figure-3) are as follows-
i) De novo fatty acid synthesis
ii) Synthesis of cholesterol
iii) Synthesis of steroids
iv) Synthesis of Sphingolipids
v) Synthesis of neurotransmitters
vi) Microsomal desaturation of fatty acids
vii) Conversion of Phenyl Alanine to Tyrosine
viii) Drug detoxification
ix) Reduction of glutathione
x) Reduction of folate
xi) Reduction of Met Hb to normal Hb
xii) The conversion of Ribonucleotides to deoxy ribonucleotides (through the action of ribonucleotide reductase) requires NADPH as the electron source; therefore, any rapidly proliferating cell needs large quantities of NADPH.
xiii) Macrophageal functions
Figure-3- NADPH is an important biological reducing agent, required for reductive biosynthesis , macrophageal function, maintenance of RBC membrane integrity and lens transparency .
2. To provide the cell with ribose-5-phosphate (R5P) for the synthesis of the nucleotides, nucleic acids ATP and coenzymes (Figure-4).
3. Although not a significant function of the PPP, it can operate to metabolize dietary pentose sugars derived from the digestion of nucleic acids as well as to rearrange the carbon skeletons of dietary pentoses into glycolytic/gluconeogenic intermediates. Glyceraldehyde-3-P and fructose-6-Pformed from 5‐C sugar phosphates may enter Glycolysis for ATP synthesis. The Pentose Phosphate Pathway thus serves as an entry into Glycolysis for both 5‐carbon & 6‐carbon sugars (Figure-4) .
4. CO2 produced from this pathway can be utilized for CO2 fixation reactions.
Figure-4- HMP pathway is an alternative pathway of glucose utilization meant for production of NADPH, required for reductive biosyntheses ; synthesis of ribose, required for the synthesis of nucleotides and nucleic acids and indirect source of energy through oxidation of glycolytic intermediates.
b) Clinical Significance
Glucose-6-phosphatase dehydrogenase (G6PD) deficiency
Glucose-6-phosphatase dehydrogenase (G6PD) deficiency is the most common disease-producing enzymopathy in humans
Inheritance– Inherited as an X-linked disorder
Frequency- Glucose-6-phosphatase dehydrogenase (G6PD) deficiency affects 400 million people worldwide.
Pathophysiology– Reactive oxygen species (ROS) generated in oxidative metabolism inflict damage on all classes of macromolecules and can ultimately lead to cell death. Indeed, ROS are implicated in a number of human diseases. Reduced glutathione (GSH), a tripeptide with a free sulfhydryl group, is required to combat oxidative stress and maintain the normal reduced state in the cell. Oxidized glutathione (GSSG) is reduced by NADPH generated by glucose 6-phosphate dehydrogenase in the pentose phosphate pathway (Figur-6).
Figure-5- NADPH produced from HMP pathway is used for reduction of oxidized glutathione that is required for the action of glutathione peroxidase enzyme , a Selenium containing metalloenzyme. In the absence of G6PD enzyme reduced availability of NADPH causes inhibition of activity of Glutathione peroxidase , which is required for the decomposition of H2O2. Accumulated H2O2 results in triggering of free radicle chain reaction and conversion of Hb to met Hb. By both these effects, life span of red blood cells is shortened.
Indeed, cells with reduced levels of glucose 6-phosphate dehydrogenase are especially sensitive to oxidative stress. This stress is most acute in red blood cells because, lacking mitochondria; they have no alternative means of generating reducing power.
G6PD deficiency is a prime example of a hemolytic anemia due to interaction between an intracorpuscular and an extracorpuscular cause, because in the majority of cases hemolysis is triggered by an exogenous agent. People deficient in glucose-6-phosphatase dehydrogenase (G6PD) are not prescribed oxidative drugs, because their red blood cells undergo rapid hemolysis under this stress. Although in G6PD-deficient subjects there is a decrease in G6PD activity in most tissues, this is less marked than in red cells, and it does not seem to produce symptoms.
Figure-6 -Red blood cells lacking G6P dehydrogenase fail to maintain RBC integrity and undergo premature lysis.
Acute HA can develop as a result of three types of triggers: (1) fava beans, (2) infections, and (3) drugs like- Antimalarials, antibiotics, Antipyretics/ analgesics, sulfonamides etc
The presence of pamaquine, a purine glycoside of fava beans, or other nonenzymatic oxidative agents leads to the generation of peroxides, reactive oxygen species that can damage membranes as well as other biomolecules. Peroxides are normally eliminated by glutathione peroxidase with the use of glutathione as a reducing agent.
Moreover, in the absence of the enzyme, the hemoglobin sulfhydryl groups can no longer be maintained in the reduced form and hemoglobin molecules then cross-link with one another to form aggregates called Heinz bodies on cell membranes. A membrane damaged by the Heinz bodies and reactive oxygen species become deformed and the cell is likely to undergo lysis. In the absence of oxidative stress, however, the deficiency is quite benign.
1) The vast majority of people with G6PD deficiency remain clinically asymptomatic throughout their lifetime.
2) However, there is an increased risk of developing neonatal jaundice (NNJ) and a risk of developing acute HA when challenged by a number of oxidative agents.
3) The onset can be extremely abrupt, especially with favism in children. The anemia is moderate to extremely severe, usually normocytic normochromic, and due partly to intravascular hemolysis; hence, it is associated with haemogobinemia and hemoglobinuria,
The laboratory workup for glucose-6-phosphate dehydrogenase (G6PD) deficiency includes the following:
1) Measurement of enzyme activity of G6PD
2) A complete blood cell (CBC) count with the reticulocyte count to determine the level of anemia and bone marrow function.
3) Indirect bilirubinemia occurs with excessive hemoglobin degradation and can produce clinical jaundice.
4) Serum Haptoglobin levels serve as an index of hemolysis and will be decreased.
5) LDH is high and so is the unconjugated bilirubin, indicating that there is also extravascular hemolysis.
Abdominal ultrasound may be useful in assessing for splenomegaly and gallstones in cases of glucose-6-phosphate dehydrogenase (G6PD) deficiency.
Peripheral Blood Film
The blood film shows anisocytosis, polychromasia, and spherocytes. The most typical feature is the presence of bizarre poikilocytes with red cells that appear to have unevenly distributed hemoglobin and red cells that appear to have had parts of them bitten away (bite cells or blister cells (Figure-7).
Figure-7- Acute hemolysis from glucose-6-phosphate dehydrogenase deficiency is linked to the development of Heinz bodies, which are composed of denatured hemoglobin
Identification and discontinuation of the precipitating agent is critical in cases of glucose-6-phosphatase dehydrogenase (G6PD) deficiency. Affected individuals are treated with oxygen and bed rest, which may afford symptomatic relief. Prevention of drug-induced hemolysis is possible in most cases by choosing alternative drugs.When acute HA develops and once its cause is recognized, no specific treatment is needed in most cases. However, if the anemia is severe, it may be a medical emergency, especially in children, requiring immediate action, including blood transfusion.
Patients must avoid broad beans (ie, fava beans). Favism occurs only in the Mediterranean variety of glucose-6-phosphate dehydrogenase (G6PD) deficiency.
Most individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency do not need treatment.Please help "Biochemistry for Medics" by CLICKING ON THE ADVERTISEMENTS above!