Metabolism of fatty acids, ketone bodies.

Questions 

01. Reaction of synthesis and utilization of ketone bodies. Biological role.

02. Mechanism of ketosis in diabetes mellitus and starvation. Ketoacidosis.

03. Biosynthesis of fatty acids:

3.1. Sources of acetyl CoA and NADPH I cytoplasm.

3.2. Synthesis of malonyl CoA.

3.3. Fatty acids synthesis, structure.

3.4. Biosynthesis of palmitic acid: reactions.

4. Biosynthesis of triacylglycerols.

5. Biosynthesis of phospholipids.

6. Fatty infiltration of liver. Lipotropic agents.

01. Reaction of synthesis and utilization of ketone bodies. Biological role.

Metabolism of ketone bodies

-         Fasting,

-         prolonged physical exertion and cases when the cells do not get enough glucose (a diet low in carbohydrates,

-         gastrointestinal disorders,

-         glucosuria, and diabetes mellitus)

activates the breakdown of fat in adipose tissue.

Fatty acids are transported in the liver in a larger amount than usually which increases speed of β-oxidation.

-         TCA cycle activity is reduced in these conditions,

-         because oxaloacetate is used in gluconeogenesis.

As a result, the rate of acetyl CoA formation exceeds the ability of TCA cycle to oxidize it.

Acetyl-CoA accumulates in the mitochondria of the liver and is used for the synthesis of acetoacetate.

-         This substance may be released into the blood by the liver or converted to other ketone body – β-hydroxybutyrate by reduction.

In hepatocyte with active β-oxidation, a high concentration of NADH occurs.

 It helps to transform mostly acetoacetate to β-hydroxybutyrate, so the main blood ketone body is β-hydroxybutyrate.

At high concentrations of acetoacetate, its part is decarboxylated non-enzymatically and turns into acetone.

-         Acetone is not utilized by tissue, but is excreted in the urine and exhaled air.

In this way the body removes excess amount of ketone bodies, which do not have time to oxidize and cause acidosis.

The rate of synthesis of ketone bodies depends on the activity of 3-hydroxy-3-methylglutaryl-CoA synthase (HMG-CoA synthase).

-         This enzyme is inducible,

-         its synthesis increases with increasing the concentration of fatty acids in the blood.

-         HMG-CoA synthase is inhibited by high concentrations of free CoA.

A small amount of ketone bodies (their concentration in the blood of 10-30 mg/liter or up to 0.2 mmol/l) is norm.

In the liver acetoacetate cannot be oxidized,

-         so, it flows with the blood into skeletal muscle, heart, brain, which is capable of converting acetoacetic acid again to acetyl-CoA.

Content of ketone bodies in the blood increases when the main source of energy for the body are fatty acids - in the prolonged muscular work, starvation, diabetes melitus.

Increase of ketone bodies concentration in the blood is called ketonemia,

the allocation of ketone bodies in the urine is called ketonuria.

Accumulation of ketone bodies in the body leads to ketoacidosis:

-         alkali reserve reduces, and in severe cases – a shift of pH occurs,

-         as β-hydroxybutyrate and acetoacetate are water-soluble organic acids capable of dissociation.

Acidosis reaches dangerous quantities in case of diabetes melitus.

The content of ketone bodies in the blood in this disease increases 100 and more times, achieving a concentration of 4-5 g/l.

Severe form of acidosis is one of the main causes of death in diabetes melitus.

In extrahepatic tissues acetoacetate is activated to acetoacetyl-CoA by succinyl-CoA-acetoacetate CoA transferase

Second mechanism

BIOMEDICAL IMPORTANCE

Fatty acids are broken down in mitochondria by oxidation to acetyl-CoA in a process that generates large amounts of energy.

When this pathway is proceeding at a high rate, three compounds,

-         acetoacetate,

-         D-3hydroxybutyrate, and

-         acetone,

known collectively as the ketone bodies, are produced by the liver.

Acetoacetate and D-3-hydroxybutyrate are used as fuels by extrahepatic tissues in normal metabolism, but overproduction of ketone bodies causes ketosis.

Increased fatty acid oxidation and consequently ketosis is a characteristic of starvation and of diabetes mellitus.

Since ketone bodies are acidic, when they are produced in excess over long periods, as in diabetes, they cause ketoacidosis, which is ultimately fatal.

Because gluconeogenesis is dependent on fatty acid oxidation, any impairment in fatty acid oxidation leads to hypoglycemia.

This occurs in various states of carnitine deficiency or deficiency of essential enzymes in fatty acid oxidation, for example, carnitine palmitoyltransferase, or inhibition of fatty acid oxidation by poisons, for example, hypoglycin.

The acetyl-CoA formed in β-oxidation is oxidized in the citric acid cycle, or it enters the pathway of ketogenesis to form ketone bodies.

As the level of serum FFA is raised, proportionately more FFA is converted to ketone bodies and less is oxidized via the citric acid cycle to CO2.

A fall in the concentration of oxaloacetate, particularly within the mitochondria, can impair the ability of the citric acid cycle to metabolize acetyl-CoA and divert fatty acid oxidation toward ketogenesis.

Such a fall may occur because of an increase in the NADH/NAD+ ratio caused by increased β-oxidation of fatty acids affecting the equilibrium between oxaloacetate and malate, leading to a decrease in the concentration of oxaloacetate,

and when gluconeogenesis is elevated, which occurs when blood glucose levels are low.

The activation of pyruvate carboxylase, which catalyzes the conversion of pyruvate to oxaloacetate partially alleviates this problem, but in conditions such as starvation and untreated diabetes mellitus, ketone bodies are overproduced causing ketosis.

Concentration of Ketone Bodies

• Concentration of total ketone bodies in the blood of wellfed individuals does not normally exceed 0.2 mmol/L (10-30 mg/L)

• Urine: Loss via urine is usually less than 1 mg/day in humans.

02. Mechanism of ketosis in diabetes mellitus and starvation. Ketoacidosis.

Ketoacidosis Results from Prolonged Ketosis Higher than normal quantities of ketone bodies present in the blood or urine constitute ketonemia (hyperketonemia) or ketonuria, respectively.

The overall condition is called ketosis.

The basic form of ketosis occurs in starvation and involves depletion of available carbohydrate coupled with mobilization of FFA.

This general pattern of metabolism is exaggerated to produce the pathologic states found in diabetes mellitus,

-         the type 2 form of which is increasingly common in Western countries;

-         twin lamb disease; and

-         ketosis in lactating cattle.

Nonpathologic forms of ketosis are found under conditions of

-         high-fat feeding and

-         after severe exercise in the postabsorptive state.

Acetoacetic and 3-hydroxybutyric acids are both moderately strong acids and are buffered when present in blood or other tissues.

However, their continual excretion in quantity progressively depletes the alkali reserve, causing ketoacidosis.

This may be fatal in uncontrolled diabetes mellitus.

03. Biosynthesis of fatty acids:

3.1. Sources of acetyl CoA and NADPH I cytoplasm.

3.2. Synthesis of malonyl CoA.

3.3. Fatty acids synthesis, structure.

3.4. Biosynthesis of palmitic acid: reactions.

There are few systems for fatty acid synthesis.

01.Extramitochondrial system: responsible for de novo synthesis of palmitic acid (always end product) from acetyl-CoA(start).

-          (De Novo Synthesis)

-         The synthesis takes place in cytosol.

02. Chain Elongation Systems:

a)     Microsomal: present in microsomes which can lengthen existing fatty acid chains.

b)     Mitochondrial: this system is mostly restricted to lengthening of an existing fatty acid of moderate chain-length.

It operates under anaerobiosis and is favored by a high NADH/NAD+ ratio.

Materials Required for the Synthesis

01. Enzymes

–        Acetyl-CoA carboxylase,

–        Fatty acid synthase, a multienzyme complex

02. Coenzymes and cofactors:

-         Biotin, NADPH, Mn++

03. CO2

04. ATP

 

3.1. Sources of acetyl CoA and NADPH I cytoplasm.

Sources of acetyl-CoA

Acetyl-CoA is mainly found in mitochondria and cannot pass out.

It forms citrate by condensing with oxaloacetate.

Citrate is transported out.

Once in cytoplasm, an enzyme citrate lyase cleaves citrate to form acetyl-CoA and oxaloacetate.

Carnitine-acetyl transferase may probably transfer acetyl group of acetyl-CoA to carnitine to form acetylcarnitine in mitochondria.

After translocation to cytoplasm acetyl group may be transferred to CoA to make it acetyl-CoA.

Sources of NADPH

Pentose phosphate pathway is the main source of NADPH.

Cytoplasmic enzyme called malic enzyme (NADP-malate dehydrogenase) catalyzed the reaction in which malate is oxidatively decarboxylated to pyruvate and NADPH is produced.

Cytoplasmic isocitrate dehydrogenase uses NADP as the coenzyme.

3.2. Synthesis of malonyl CoA.

Reaction occurs in two steps:

01. Biotin-enzyme + ATP + HCO3 ^–

Carboxy-biotin-enzyme +ADP +Pi

 

02.Carboxy-biotin-enzyme + Acetyl-CoA

 

      Malonyl CoA + biotin-enzyme

 

The reaction is irreversible.

Regulation

Acetyl-CoA carboxylase catalyzes the rate limiting step in the de novo synthesis of fatty acids and provides the earliest point at which control can be exerted.

The enzyme is inactivated by phosphorylation.

A:  Insulin, CoA, Guanine nucleotides

I:  Glucagon, adrenaline, acyl-CoA Decrease in citrate concentration decreases acetyl-CoA carboxylase activity

 

 

3.3. Fatty acids synthase, structure.

It is a multienzyme complex.

It is made up of an ellipsoid dimer of two identical polypeptide monomeric units,

arranged in a “head to tail” fashion

The ACP has an –SH group in the 4phosphopantothene moiety, referred as pantothenyl-SH (Pan-SH)

Another active –SH group present in the cysteine moiety of the enzyme ketoacyl synthase, referred as cysteinyl-SH (Cys–SH).

The “Pan-SH” of one monomeric unit is in close proximity to the “Cys-SH” group of other monomeric unit and vice-versa.

Sequence of domaines in primary structure of fatty acid syhthase monomer

-         ketoacyl synthase,

-         malonyl/acetyl transacylase,

-         hydratase

-         enoyl reductase,

-         ketoacyl reductase,

-         ACP

-         thio-esterase (deacylase)

Complex is functional only when the two monomeric units are in association with each other.

The functional activity is lost when they are dissociated.

In a dimer form, the complex jointly synthesises 2 molecules of palmitic acid simultaneously.

Sequence of enzyme domains in primary structure of fatty acid synthase monomer

3.4. Biosynthesis of palmitic acid: reactions.

4. Biosynthesis of triacylglycerols.

TAG (neutral fats) is the main form of energy deposition.

Deposited fat can provide the body with energy during fasting for a long time (up to 7-8 weeks).

TAG synthesis occurs in absorptive period in the liver and adipose tissue.

-         However, if the adipose tissue participates only in fat deposition,

-         the liver plays an important role in converting carbohydrates originating from food in fats which are then secreted into the blood as part of VLDL and delivered to other tissues.

The immediate substrates for the synthesis of fats are the acyl- CoA and glycerol-3-phosphate.

The metabolic pathway of synthesis of fats in the liver and adipose tissue is the same,

-         except for the different pathways of glycerol-3-phosphate synthesis.

The liver is the main organ where synthesis of fatty acids from the products of glycolysis takes place.

In the smooth endoplasmic reticulum of hepatocytes fatty acids, interacting with glycerol-3phosphate, are activated and immediately used for the synthesis of TAG.

Synthesised fats are packaged in VLDL and secreted into the blood.

In adipose tissue for TAG synthesis mainly fatty acids released by the hydrolysis of ChM and VLDL fats, are used.

Fatty acids come into adipocytes, where they are transformed into derivatives of CoA and react with glycerol-3-phosphate.

Furthermore, in these cells synthesis of fatty acids from products of glycolysis occurs.

TAG molecules in adipocytes are combined into larger oil droplets, containing no water, which is the most compact form of fuel storage molecules.

5. Biosynthesis of phospholipids.

Phospholipids

Phospholipids are a specialized group of lipids performing a variety of functions.

These include the membrane structure & functions   involvement in blood clotting &

supply of arachidonic acid for the synthesis of prostaglandins.

Synthesis of phospholipids

Phospholipids are synthesized from phosphatidic acid & 1.2-diacylglycerol,

intermediates in the production of triacylglycerols.

Phospholipid’s synthesis occurs in the smooth endoplasmic reticulum.

Inner mitochondrial membrane

 

Formation of lecithin & cephalin

It occurs mainly in liver & brain.

Choline & ethanolamine first get phosphorylated & then combine with CTP to form CDP-choline & CDP-ethanolamine.

Phosphatidylcholine (lecithin) is synthesized when CDP-choline combines with 1,2-diacylglycerol.

 

Synthesis of phosphatidylserine

Phosphatidyl ethanolamine can exchange its ethanolamine group with free serine to produce phosphatidylserine.

 

Formation of phosphatidylinositol

CDP-diacylglycerol produced from phosphatidic acid combines with inositol to form phosphatidyl inositol (PI).

Phosphatidyl inositol contains arachidonic acid on carbon 2of glycerol which serves as a substrate for prostaglandin synthesis.

Pl is important for signal transmission across membranes.

 

The synthesis of phospholipids requires

• glycerol

• fatty acids

• inorganic phosphates

• nitrogen bases (in particular, choline for synthesis of phosphatidylcholine)

6. Fatty infiltration of liver. Lipotropic agents.

-         In insufficient synthesis of choline, or its short supply to the liver,

-         the synthesis of phospholipids from neutral fat components becomes either impossible, or drastically decreased,

which results in deposition of neutral fats in the liver.

Such condition is referred to as fatty infiltration of liver,

which may subsequently develop into a fatty degeneration of the liver (steatosis)

In other words, the synthesis of phospholipids needs either choline or compounds capable to act as methyl group donors and thus participate in the production of choline (for example, methionine)

Such compounds are known as lipotropic agents

curd cheese is recommended in the diet as lipotropic agent,

since its ingredient is casein, a protein whose molecule contains a large number of methionine residues