Glycolysis

we study carbohydrate digestion and know that the end of digestion is production of Glucose, Fructose and Galactose

Also we know that glucose enters the liver through the hepatic portal vein and glucose should be oxidized through glycolysis

Glycolysis

A)  Definition:

It is the breakdown of glucose in the cell cytosol (= cytoplasm) producing pyrurate in the presence of oxygen or lactate in the absence of oxygen.

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B)   Site:

Glycolysis occurs in cytoplasm but:

• In presence of O₂, oxidation of glucose is complete in mitochondria where pyruvate enters the kerb’s cycle and the electron transport chain to complete the oxidation of glucose resulting in a high amount of energy.

• In absence of O­₂, pyruvate is converted into lactate in the cytoplasm giving a small amount of energy but it is important to some tissue.

Therefore:

Occurrence of glycolysis is of physiological importance in:

1. Tissues with no mitochondria such as RBCs, cornea and lens.

2. Tissues with few mitochondria: Testis, leucocytes, medulla of the kidney, retina, skin and gastrointestinal tract

3. Tissues undergo frequent oxygen lack: skeletal muscles especially during exercise.

Where they depend only on the glycolysis not on kerbs cycle and the electron transport chain

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C)    Stages of glycolysis:

<<<<<<<Stage (I)>>>>>>>

It is the energy requiring stage

in this stage:

• One molecule of glucose is converted into two molecules of glyceraldehyde-3-phosphate.

• These steps consume 2 molecules of ATP .

Step one:

1.      Event: (glucose phosphorylation)­

===> A phosphate group is transferred from ATP molecule to the carbon number 6 in the glucose molecule forming glucose-6-phosphate, thus this step is energy consuming.

===> Glucose-6-phosphate is an intermediate forming an important branch point in the metabolism.

===> This step is fast irreversible step

2.  Enzymes stimulating this step:

===> The enzymes used are either glucokinase or hexokinase enzymes which are responsible for entry of glucose into the cell and phosphorylation of glucose which lead to glucose trapping inside the cell, therefore this step is irreversible because if it is reversible the glucose-6-ph will return to glucose and could exit from the cell again.

===>They are activated after a carbohydrate rich meal for 2 hours to lower the glucose blood level.

NOTE: Kinase enzyme always add phosphate group on the substrate

What is the difference between Hexokinase and glucokinase? (click to open)

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ٍStep 2:

1.   Event: (formation of fructose-6-phosphate from glucose-6-phosphate)­

Isomerization of glucose-6-phosphate to fructose 6-phosphate, I.e. a conversion of an aldose into a ketose by phosphoglucose isomerase.

It doesn’t need energy because it occurs spontaneosly.

2.   Enzymes stimulating this step: phosphoglucose isomerase.

NOTE: Isomerase enzyme always catalyzes the structural rearrangements.

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Step 3:

a.  Event: (Phosphorylation of Fructose-6-phosphate to fructose-1,6-­bisphosphate)­

-  Phosphorylation of Fructose-6-phosphate by ATP to fructose-1,6-­bisphosphate (F-1 ,6-BP)

-   This step is irreversible.

b.  Enzymes stimulating this step: by phosphofructokinase (PFK)

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step 4:

a. Event: (cleave of 6-carbon sugar into two 3-carbon fragments)­

-  Splitting of fructose-1,6-bisphosphate into tow 3-carbon fragments:

• Glyceraldehyde 3-phosphate (GAP)
• Dihydroxyacetone phosphate (DHAP)

-   Reversible under intracellular conditions

b.  Enzymes stimulating this step:

-  Aldolase. (This enzyme derives its name from the nature of the reverse reaction, an aldol condensation).

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Step 5:

a.  Event: (isomerisation of (GAP) to (DHAP))­

-  Isomerisation of Glyceraldehyde 3-phosphate (GAP) to Dihydroxyacetone phosphate (DHAP)

-  It is fast reversible step.

b. Enzymes stimulating this step: triose phosphate isomerase (TPI or TIM).

c.  Importance of this step:

To get full energy from glucose molecules in other words to get energy from the 6 carbons. EXPLAIN?

-   The cell can’t oxidize DHAP to get energy from it , thus it will be lost as a waste product.

-    Thus, the cell will obtain energy from GAP only (i.e. from 3 carbon of the glucose), not from the 6-carbons.

-    But, the GAP will continue the glycolysis and give energy, while DAHP is not

-     Thus, in order to get energy from the 6-carbons of glucose, the DHAP must be converted to GAP

After this step we get 2 molecules of Glyceraldehyde 3-phosphate (GAP):

One form step 4 and one form step 5.

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<<<<<Stage (II)>>>>>

(The energy producing stage)

In this stage:

Tow moleculse of Glyceraldehyde 3-phosphate (GAP) is converted into two molecules of pyruvate producing 4 molecules of ATP.

 

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Step 6

a.  Event: (Oxidation of glyceraldehyde 3-phosphate to 1,3-bisphosphoglycerate)

Conversion of glyceraldehyde-3-phosphate into 1,3-­bisphosphoglycerate (1,3-BPG), a reaction catalyzed by glyceraldehyde-3-phosphate dehydrogenase.

b.  Enzymes stimulating this step: glyceraldehyde 3-phosphate dehydrogenase.

Note that, the phosphate used in this step is inorganic phosphate.

The oxidation of the aldehyde to an acid is coupled to the reduction of NAD⁺ to NADH/H⁺

Importance of 1,3-bisphosphoglycerate:

- During the glycolysis inside the RBCs, part of the 1,3-bisphosphoglycerate is converted into 2,3-bisphosphoglycerate by the enzyme bisphosphoglycerate mutase

-   2,3-BPG decreases the affinity of Haemoglobin for Oxygen (i.e. decreases the attachment of O₂ to haemoglobin) thus the oxygen can leave hemoglobin easily and this is good in case of oxygen shortage because cells will be able to get their needs of O₂ easily even if the oxygen supply is low.

Clinical and physiological aspects of 2,3-BPG تطبيقات للكلام اللى قولناه

1. smokers and people who live in high altitude, where the Hb increase in each RBC and the number of RBCs increase and increase the amount of 2,3-bisphosphoglycerate leading to decrease the affinity of Hb for O₂ causing dissociation of O₂ form Hb easily into the blood capillary and thus the cells takes its need easily.

2. Fetus gets oxygen form the mother thus fetal Hb has a high affinity for O₂because it has a plenty of O₂ from his mother, thus competing with the Hb of the mother because the oxygen leave the Mother Hb because of it low affinity and attach to the Fetus Hb which has a high affinity for the O₂.

3. The sotred blood for blood transfusion has diminished levels of 2,3-BPG, thus before transfusion of inosine or the glycolytic substrate dihydroxyphosphate to regenerate 2,3-BPG before blood transfusion because in absence of 2,3-BPG suffocation اختناق occurs.

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step 7:

a.  Event: (Conversion of 1,3-bisphosphoglycerate to 3­phosphoglycerate)

-  Conversion of 1,3-bisphosphoglycerate to 3-­phosphoglycerate

-  This step gproduce one ATP molecule.

b.  Enzymes stimulating this step: phosphoglycerate kinase.

Note that:

Formation of ATP from transferring a Phosphate group to ADP from organic substrate is called Substrate level phosphorylation.

Formation of ATP from by transferring a Phosphate group to ADP from NAD or FAD in the electron transport chain is called Oxidative phosphorylation.

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Step 8:

a.   Event: (Conversion of 3-phosphoglycerate to 2­-phosphoglycerate)

Shifting the phosphate group from the carbon number 3 to the carbon number 2

a.   Enzymes stimulating this step: phosphoglycerate mutase.

The importance of this step is to make the compound suitable for the active site of the enolase enzyme.

NOTE: Mutase shifting of a functional group from one position to another within the same molecule

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ٍStep 9:

a.  Event: (2-Phosphoglycerate is dehydrated to form phosphoenolpyruvate)

The dehydration of the alcohol produces a double bond between carbons 2 and 3 and creates a high-energy enol phosphate linkage.

b.  Enzymes stimulating this step: enolase.

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Step 10:

a.  Event: (Conversion of phosphoenol pyruvate to pyruvate)

Conversion of phosphoenol pyruvate to pyruvate producing one ATP molecule.

It is irreversible step.

b.  Enzymes Stimulating this step: Pyruvate Kinase (transfer Phosphate group from phosphoenolo-pyruvate to ADP molecule to produce one ATP Molecule)

فالخطوة دى برضو اتكون عندنا kinase يعنى هينقل مجموعة فوسفات وبالفعل هو نقل مجموعة فوسفات من الphosphoenolo-pyruvate الى جزئ ADP عشان يدينا جزء واحد من الATP.

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D)  Calculation of ATP molecules from glycolysis:

We know that we get from STAGE (I) 2 molecules of glyceraldehyde-3-phosphate and we know that each molecules when continue glycolysis in the STAGE(II) give 2 ATP molecule, thus the 2 molecule of glyceraldehyde-3-phosphate give 4 ATP molecules.

Also we know that the steps number 1 and 3 consume 2 molecule of ATP

Thus, the net product of ATP during glycolysis is 2 ATP

4 ATP form Stage (II) – 2 ATP from Stage (I) = 2 ATP

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E)    Regulation of Glycolysis:

Glycolysis pathway is regulated by:

A) Control of 3 enzymes that catalyze the 3 irreversible steps of glycolysis:

1. Hexokinase (step 1),

2. Phosphofructokinase (step 3)

3. Pyruvate Kinase (step 10).

B) Energy regulation:

1. High level of ATP inhibits  phosphofructokinase (PFK-1) and pyruvate kinase.

2. High level of ADP and AMP stimulate PFK.

C) Substrate regulation:

1.  Glucose-6-phosphate inhibits hexokinase (and not glucokinase).

2. Fructose 1,6 bisphosphate stimulates phosphofructokinase-1.

3. Citrate inhibits phosphofructokinase-1.

4. Fructose 1,6 bisphosphate stimulates pyruvate kinase.

D) Hormonal regulation:

1. Insulin: Stimulates synthesis of all key enzymes of glycolysis. It is secreted after meal (in response to high blood glucose level).

2. Glucagon: Inhibits the activity of all key enzymes of glycolysis. It is secreted in response to low blood glucose level.

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Diabetes Mellitus- Etiology of Type 2 and Pathophysiology of Type 1 Diabetes mellitus

Etiology of Type 2 Diabetes Mellitus

This represents a heterogeneous group of conditions that used to occur predominantly in adults, but it is now more frequently encountered in children and adolescents. More than 90% of all diabetic persons in the United States are included under this classification. Circulating endogenous insulin is sufficient to prevent ketoacidosis but is inadequate to prevent hyperglycemia in the face of increased needs owing to tissue insensitivity (insulin resistance).

Genetic considerations

Genetic and environmental factors combine to cause both the insulin resistance and the beta cell loss. Most epidemiologic data indicate strong genetic influences, since in monozygotic twins over 40 years of age, concordance develops in over 70% of cases within a year whenever type 2 diabetes develops in one twin. Individuals with a parent with type 2 DM have an increased risk of diabetes; if both parents have type 2 DM, the risk approaches 40%. Insulin resistance, as demonstrated by reduced glucose utilization in skeletal muscle, is present in many nondiabetic, first-degree relatives of individuals with type 2 DM. The disease is polygenic and multifactorial since in addition to genetic susceptibility, environmental factors (such as obesity, nutrition, and physical activity) modulate the phenotype. The mechanisms by which these genetic alterations increase the susceptibility to type 2 diabetes are not clear.

Environmental factors

Obesity is the most important environmental factor causing insulin resistance. The degree and prevalence of obesity varies among different racial groups with type 2 diabetes. Visceral obesity, due to accumulation of fat in the omental and mesenteric regions, correlates with insulin resistance; subcutaneous abdominal fat seems to have less of an association with insulin insensitivity. Exercise may affect the deposition of visceral fat as suggested by CT scans of Japanese wrestlers, whose extreme obesity is predominantly subcutaneous. Their daily vigorous exercise program prevents accumulation of visceral fat, and they have normal serum lipids and euglycemia despite daily intakes of 5000–7000 kcal and development of massive subcutaneous obesity.

Several adipokines, secreted by fat cells, can affect insulin action in obesity. Two of these, leptin and adiponectin, seem to increase sensitivity to insulin, presumably by increasing hepatic responsiveness.Two others—tumor necrosis factor-α, which inactivates insulin receptors, and the newly discovered peptide, resistin—interfere with insulin action on glucose metabolism and have been reported to be elevated in obese animal models. Mutations or abnormal levels of these adipokines may contribute to the development of insulin resistance in human obesity.

Other Specific Types of Diabetes Mellitus

Maturity-onset diabetes of the young (MODY)

This subgroup is a relatively rare monogenic disorder characterized by non–insulin-dependent diabetes with autosomal dominant inheritance and an age at onset of 25 years or younger. Patients are nonobese, and their hyperglycemia is due to impaired glucose-induced secretion of insulin. Six types of MODY have been described. Except for MODY 2, in which a Glucokinase gene is defective, all other types involve mutations of a nuclear transcription factor that regulates islet gene expression.

MODY 2 is quite mild, associated with only slight fasting hyperglycemia and few if any microvascular diabetic complications. It generally responds well to dietary modifications or low doses of oral hypoglycemic agents. MODY 3—the most common form—accounts for two-thirds of all MODY cases. The clinical course is similar to that of idiopathic type 2 diabetes in terms of microangiopathy and failure to respond to oral agents with time.

Diabetes due to mutant insulins

This is a very rare subtype of nonobese type 2 diabetes, with no more than ten families having been described. Since affected individuals were heterozygous and possessed one normal insulin gene, diabetes was mild, did not appear until middle age, and showed autosomal dominant genetic transmission. There is generally no evidence of clinical insulin resistance, and these patients respond well to standard therapy.

Diabetes due to mutant insulin receptors

Defects in one of their insulin receptor genes have been found in more than 40 people with diabetes, and most have extreme insulin resistance associated with acanthosis nigricans. In very rare instances when both insulin receptor genes are abnormal, newborns present with a leprechaun-like phenotype and seldom live through infancy.

Diabetes mellitus associated with a mutation of mitochondrial DNA

Since sperm do not contain mitochondria, only the mother transmits mitochondrial genes to her offspring. Diabetes due to a mutation of mitochondrial DNA that impairs the transfer of Leucine or lysine into mitochondrial proteins has been described. Most patients have a mild form of diabetes that responds to oral hypoglycemic agents; some have a nonimmune form of type 1 diabetes. Two-thirds of patients with this subtype of diabetes have a hearing loss, and a smaller proportion (15%) had a syndrome of myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS).

Wolfram's syndrome

Wolfram's syndrome is an autosomal recessive neurodegenerative disorder first evident in childhood. It consists of diabetes insipidus, diabetes mellitus, optic atrophy, and deafness, hence the acronym DIDMOAD. It is due to mutations in a gene named WFS1, which encodes a 100.3 KDa transmembrane protein localized in the endoplasmic reticulum. The function of the protein is not known.

Transient or permanent neonatal diabetes

Onset < 6 months of age, may be caused by several genetic mutations and requires treatment with insulin. Mutations in subunits of the ATP-sensitive potassium channel subunits are the major causes of permanent neonatal diabetes. Although these activating mutations impair glucose-stimulated insulin secretion, these individuals may respond to sulfonylureas and improve their glycemic control and can be treated with these agents. Homozygous Glucokinase mutations cause a severe form of neonatal diabetes.

Insulin Resistance Syndrome (Syndrome X; Metabolic Syndrome)

Twenty-five percent of the general nondiabetic obese population has insulin resistance of a magnitude similar to that seen in type 2 diabetes. These insulin-resistant nondiabetic individuals are at much higher risk for developing type 2 diabetes than insulin-sensitive persons. In addition to diabetes, these individuals have increased risk for elevated plasma triglycerides, lower high-density lipoproteins (HDLs), and higher blood pressure—a cluster of abnormalities termed syndrome X. These associations have now been expanded to include small, dense, low-density lipoprotein (LDL), hyperuricemia, abdominal obesity, prothrombotic state with increased levels of plasminogen activator inhibitor type 1 (PAI-1), and proinflammatory state.These clusters of abnormalities significantly increase the risk of atherosclerotic disease.

Pathophysiology of Type 1 Diabetes

Although other islet cell types [alpha cells (glucagon-producing), delta cells (somatostatin-producing), or PP cells (pancreatic polypeptide-producing)] are functionally and embryologically similar to beta cells and express most of the same proteins as beta cells, they are inexplicably spared from the autoimmune process.

Pathologically, the pancreatic islets are infiltrated with lymphocytes (in a process termed insulitis). After all beta cells are destroyed, the inflammatory process abates, the islets become atrophic, and most immunologic markers disappear. The precise mechanisms of beta cell death are not known but may involve the formation of nitric oxide metabolites, apoptosis, and direct CD8+ T cell cytotoxicity. The autoimmune destruction of pancreatic β-cells leads to a deficiency of insulin secretion. It is this loss of insulin secretion that leads to the metabolic derangements associated with IDDM.

In addition to the loss of insulin secretion, the function of pancreatic α-cells is also abnormal. There is excessive secretion of glucagon in IDDM patients. Normally, hyperglycemia leads to reduced glucagon secretion. However, in patients with IDDM, glucagon secretion is not suppressed by hyperglycemia. The resultant inappropriately elevated glucagon levels exacerbate the metabolic defects due to insulin deficiency .The most pronounced example of this metabolic disruption is that patients with IDDM rapidly develop diabetic ketoacidosis in the absence of insulin administration. Particularly problematic for long term IDDM patients is an impaired ability to secrete glucagon in response to hypoglycemia. This leads to potentially fatal hypoglycemia in response to insulin treatment in these patients.

Although insulin deficiency is the primary defect in IDDM, in patients with poorly controlled IDDM there is also a defect in the ability of target tissues to respond to the administration of insulin. There are multiple biochemical mechanisms that account for this impairment of tissues to respond to insulin. Deficiency in insulin leads to elevated levels of free fatty acids in the plasma as a result of uncontrolled lipolysis in adipose tissue. Free fatty acids suppress glucose metabolism in peripheral tissues such as skeletal muscle. This impairs the action of insulin in these tissues, i.e. the promotion of glucose utilization.

Additionally, insulin deficiency decreases the expression of a number of genes necessary for target tissues to respond normally to insulin such as Glucokinase in liver and the GLUT 4 class of glucose transporters in adipose tissue. The major metabolic derangements which result from insulin deficiency in IDDM are impaired glucose, lipid and protein metabolism.

Glucose Metabolism: Uncontrolled IDDM leads to increased hepatic glucose output. First, liver glycogen stores are mobilized then hepatic gluconeogenesis is used to produce glucose. Insulin deficiency also impairs non-hepatic tissue utilization of glucose. In particular in adipose tissue and skeletal muscle, insulin stimulates glucose uptake. This is accomplished by insulin-mediated movement of glucose transporter proteins to the plasma membrane of these tissues. Reduced glucose uptake by peripheral tissues in turn leads to a reduced rate of glucose metabolism. In addition, the level of hepatic Glucokinase is regulated by insulin. Therefore, a reduced rate of glucose phosphorylation in hepatocytes leads to increased delivery to the blood. Other enzymes involved in anabolic metabolism of glucose are affected by insulin (primarily through covalent modifications). The combination of increased hepatic glucose production and reduced peripheral tissues metabolism leads to elevated plasma glucose levels. When the capacity of the kidneys to absorb glucose is surpassed, Glycosuria ensues. Glucose is an osmotic diuretic and an increase in renal loss of glucose is accompanied by loss of water and electrolytes, termed polyuria. The result of the loss of water (and overall volume) leads to the activation of the thirst mechanism (polydipsia). The negative caloric balance which results from the glucosuria and tissue catabolism leads to an increase in appetite and food intake (polyphagia).

Lipid Metabolism: One major role of insulin is to stimulate the storage of food energy following the consumption of a meal. This energy storage is in the form of glycogen in hepatocytes and skeletal muscle. Additionally, insulin stimulates hepatocytes to synthesize triglycerides and storage of triglycerides in adipose tissue. In opposition to increased adipocyte storage of triglycerides is insulin-mediated inhibition of lipolysis. In uncontrolled IDDM there is a rapid mobilization of triglycerides leading to increased levels of plasma free fatty acids. The free fatty acids are taken up by numerous tissues (however, not the brain) and metabolized to provide energy. Free fatty acids are also taken up by the liver.

Normally, the levels of malonyl-CoA are high in the presence of insulin. These high levels of malonyl-CoA inhibit carnitine palmitoyl Transferase I, the enzyme required for the transport of fatty acyl-CoA's into the mitochondria where they are subject to oxidation for energy production. Thus, in the absence of insulin, malonyl-CoA levels fall and transport of fatty acyl-CoA's into the mitochondria increases. Mitochondrial oxidation of fatty acids generates acetyl-CoA which can be further oxidized in the TCA cycle. However, in hepatocytes the majority of the acetyl-CoA is not oxidized by the TCA cycle but is metabolized into the ketone bodies, Acetoacetate and β-hydroxybutyrate. These ketone bodies leave the liver and are used for energy production by the brain, heart and skeletal muscle. In IDDM, the increased availability of free fatty acids and ketone bodies exacerbates the reduced utilization of glucose furthering the ensuing hyperglycemia. Production of ketone bodies, in excess of the body’s ability to utilize them leads to ketoacidosis. In diabetics, this can be easily diagnosed by smelling the breath. A spontaneous breakdown product of acetoacetate is acetone which is volatilized by the lungs producing a distinctive odor.

Normally, plasma triglycerides are acted upon by lipoprotein lipase (LPL), an enzyme on the surface of the endothelial cells lining the vessels. In particular, LPL activity allows fatty acids to be taken from circulating triglycerides for storage in adipocytes. The activity of LPL requires insulin and in its absence a hypertriglyceridemia results.

Protein Metabolism: Insulin regulates the synthesis of many genes, either positively or negatively that then affect overall metabolism. Insulin has a global effect on protein metabolism, increasing the rate of protein synthesis and decreasing the rate of protein degradation. Thus, insulin deficiency will lead to increased catabolism of protein. The increased rate of proteolysis leads to elevated concentrations in plasma amino acids. These amino acids serve as precursors for hepatic and renal gluconeogensis. In liver, the increased gluconeogenesis further contributes to the hyperglycemia seen in IDDM.

------------------------------------------ Best Wishes: Dr.Ehab Aboueladab, Tel:01007834123 Email:ehab10f@gmail.com,ehababoueladab@yahoo.com ------------------------------------------

مؤتمر صحفي للحكومة الليبية حول مقتل السفير الامريكي

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فيديو للسفير الامريكي في ليبيا كريس ستيفنز قبل مقتله

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