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Fatty acid Biosynthesis- Subjective Questions- Set-1
Q.- Discuss the important differences between fatty acid biosynthesis and fatty acid oxidation.
Answer-Fatty acid synthesis is not simply a reversal of the degradative pathway. Rather, it consists of a new set of reactions, again exemplifying the principle that synthetic and degradative pathways are almost always distinct. Some important differences between the pathways are:
1. Synthesis takes place in the cytosol, in contrast with degradation, which takes place primarily in the mitochondrial matrix.
2. Intermediates in fatty acid synthesis are covalently linked to the sulfhydryl groups of an acyl carrier protein (ACP), whereas intermediates in fatty acid breakdown are covalently attached to the sulfhydryl group of coenzyme A.
3. The enzymes of fatty acid synthesis in higher organisms are joined in a single polypeptide chain called fatty acid synthase. In contrast, the degradative enzymes do not seem to be associated.
4. The growing fatty acid chain is elongated by the sequential addition of two-carbon units derived from acetyl CoA. The activated donor of two carbon units in the elongation step is malonyl ACP. The elongation reaction is driven by the release of CO2.
5. The reductant in fatty acid synthesis is NADPH, whereas the oxidants in fatty acid degradation are NAD + and FAD.
6. Elongation by the fatty acid synthase complex stops on formation of palmitate (C16). Further elongation and the insertion of double bonds are carried out by other enzyme systems.
Q. Acetyl co A, the precursor of fatty acids is produced in the mitochondrial matrix, but the enzymes of de novo fatty acid synthesis are extra mitochondrial, How is Acetyl co A transported out of the selectively permeable mitochondrial membrane.
Answer- Fatty acids are synthesized in the cytosol, whereas acetyl CoA is formed from pyruvate in mitochondria. Hence, acetyl CoA must be transferred from mitochondria to the cytosol. Mitochondria, however, are not readily permeable to acetyl CoA. Carnitine carries only long-chain fatty acids. The barrier to acetyl CoA is bypassed by citrate, which carries acetyl groups across the inner mitochondrial membrane. Citrate is formed in the mitochondrial matrix by the condensation of acetyl CoA with oxaloacetate (Figure-1 ).
Figure-1- showing the transport of Acetyl co A in the form of citrate from mitochondrial matrix to cytosol
When present at high levels, citrate is transported to the cytosol, where it is cleaved by ATP-citrate Lyase, which increases in activity in the well-fed state. The acetyl-CoA is then available for malonyl-CoA formation and synthesis to palmitate (Figure-1).
Oxaloacetate formed in the transfer of acetyl groups to the cytosol must now be returned to the mitochondria.The inner mitochondrial membrane is impermeable to oxaloacetate. Hence, a series of bypass reactions are needed. Most importantly, these reactions generate much of the NADPH needed for fatty acid synthesis.
First, oxaloacetate is reduced to malate by NADH. This reaction is catalyzed by a malate dehydrogenase in the cytosol.
Second, malate is oxidatively decarboxylated by an NADP + -linked malate enzyme (also called malic enzyme).
The pyruvate formed in this reaction readily enters mitochondria, where it is carboxylated to oxaloacetate by pyruvate carboxylase.
The sum of these three reactions is-
Thus, one molecule of NADPH is generated for each molecule of acetyl CoA that is transferred from mitochondria to the cytosol. Hence, eight molecules of NADPH are formed when eight molecules of acetyl CoA are transferred to the cytosol for the synthesis of palmitate. The additional six molecules of NADPH required for this process come from the pentose phosphate pathway.
Alternatively, malate itself can be transported into the mitochondrion, where it is able to re-form oxaloacetate. Note that the citrate (tricarboxylate) transporter in the mitochondrial membrane requires malate to exchange with citrate (Figure -1).
Q- Discuss the central role of Acetyl co A in the metabolic pathways.
Answer- Acetyl CoA acts either as a metabolic substrate or product for various classes of biomolecules and as a major source of useful metabolic energy.
Sources of Acetyl co A- Acetyl co A, is produced from pyruvate, ketogenic amino acids, fatty acid oxidation, ketolysis and by alcohol metabolism (Figure-2).
Fate of Acetyl co A
It is a substrate for TCA cycle and a precursor for fatty acids, cholesterol, ketone bodies and steroids. It is also required for detoxification reactions and for the synthesis of acetyl choline. Acetyl CoA can react “reversibly” in the degradation or synthesis of lipids and amino acids. This is not the case with carbohydrate metabolism. In mammals, it is impossible to use acetyl CoA to make carbohydrates. Acetyl co A forms the basis for the synthesis of steroids. Some steroids of importance include cholesterol, bile salts, sex hormones, aldosterone, and cortisol.
Figure- 2-Showing the central role of Acetyl co A
Q- Give a brief account of the sources of NADPH
Answer- Sources of NADPH- NADPH is involved as donor of reducing equivalents.
The oxidative reactions of the pentose phosphate pathway (Figure-3) are the chief source of the hydrogen required for the reductive synthesis of fatty acids. Significantly, tissues specializing in active lipogenesis—ie, liver, adipose tissue, and the lactating mammary gland—also possess an active pentose phosphate pathway. Moreover, both metabolic pathways are found in the cytosol of the cell, so there are no membranes or permeability barriers against the transfer of NADPH.
Figure-3- Showing the formation of NADPH in the Pentose phosphate pathway. Per Glucose-6-P two molecules of NADPH are produced.
Other sources of NADPH include-
1) From Malate-The reaction that converts malate to pyruvate catalyzed by the “malic enzyme” (NADP malate dehydrogenase) is also an alternative source of NADPH (Figure-4)
Figure-4 – Showing the formation of NADPH from Malate by the activity of Malic enzyme
2) From Isocitrate- The extra mitochondrial isocitrate dehydrogenase reaction though not a common source but does contribute to the formation of NADPH (Figure-5). There are three isoenzymes of isocitrate dehydrogenase. One, which uses NAD+, is found only in mitochondria. The other two use NADP+ and are found in mitochondria and the cytosol. Respiratory chain-linked oxidation of isocitrate proceeds almost completely through the NAD+-dependent enzyme.
Figure- 5- Showing the formation of NADPH from the activity of cytosolic isocitrate dehydrogenase
Q- Explain briefly about the reaction catalyzed by Acetyl co A carboxylase, What is the significance of this reaction?
Answer-The synthesis of malonyl CoA is catalyzed by acetyl CoA carboxylase. Fatty acid synthesis starts with the carboxylation of acetyl CoA to malonyl CoA. This irreversible reaction is the committed step in fatty acid synthesis.
Figure- 6- Showing the formation of Malonyl co A from Acetyl co A
Acetyl-CoA carboxylase has a requirement for the vitamin biotin (Figure-7)
Figure-7- Showing the attachment of Biotin to the enzyme and the formation of carboxy biotin. Biotin is the first acceptor of the carboxyl group
The enzyme is a multienzyme protein containing a variable number of identical subunits, each containing-
2) Biotin carboxylase,
3) Biotin carboxyl carrier protein
5) A regulatory allosteric site.
Bicarbonate as a source of CO2 is required in the initial reaction for the carboxylation of acetyl-CoA to malonyl-CoA in the presence of ATP and acetyl-CoA carboxylase.
The reaction takes place in two steps: (1) carboxylation of biotin involving ATP and (2) transfer of the carboxyl to acetyl-CoA to form malonyl-CoA (Figure-8)
Figure-8 – Showing the role of biotin in the carboxylation of Acetyl co A
The first reaction which includes the carboxylation of biotin to form carboxy biotin is catalyzed with the biotin subunit of acetyl-CoA carboxylase. This portion of the mechanism is ATP dependent; also the bicarbonate provides the CO2. The second step of this mechanism requires that the carboxyl group be transferred from the biotin to the acetyl-CoA to form malonyl-CoA. This reaction thermodynamically should not be spontaneous, but because it is coupled with the hydrolysis of ATP to ADP this reaction goes. This hydrolysis of ATP also is one main reason this reaction is the committed step of this metabolic cycle. This mechanism is very much like the pyruvate kinase that is a major committed step in glycolysis that converts phosphoenolpyruvate to pyruvate.
Acetyl co A carboxylase is the rate controlling enzyme in the pathway of lipogenesis. It is regulated by-
1) Allosteric modification- Acetyl-CoA carboxylase is an allosteric enzyme and is activated by citrate, which increases in concentration in the well-fed state and is an indicator of a plentiful supply of acetyl-CoA. Citrate converts the enzyme from an inactive dimer to an active polymeric form, with a molecular mass of several million. Inactivation is promoted by long-chain acyl-CoA molecules.
2) Feedback Inhibition- The enzyme is inhibited by malonyl co A and Palmitoyl co A, an example of negative feedback inhibition by a product of a reaction.Thus, if acyl-CoA accumulates because it is not esterified quickly enough or because of increased lipolysis or an influx of free fatty acids into the tissue, it will automatically reduce the synthesis of new fatty acid. Acyl-CoA also inhibits the mitochondrial tricarboxylate transporter, thus preventing activation of the enzyme by egress of citrate from the mitochondria into the cytosol.
3) Covalent Modification-Acetyl-CoA carboxylase is also regulated by hormones such as glucagon, epinephrine, and insulin via changes in its phosphorylation state (Figure-8)
Figure-8- Regulation of acetyl-CoA carboxylase by phosphorylation/dephosphorylation, the enzyme is inactivated by phosphorylation by AMP-activated protein kinase (AMPK) . Glucagon (and epinephrine) increase cAMP, and thus activate this latter enzyme via cAMP-dependent protein kinase. Insulin activates acetyl-CoA carboxylase by causing dephosphorylation mediated by protein phosphatase.
4) Induction and Repression- Insulin is an important hormone causing gene expression and induction of enzyme biosynthesis, and glucagon (via cAMP) antagonizes this effect. Feeding fats containing polyunsaturated fatty acids coordinately regulates the inhibition of expression of key enzymes of glycolysis and lipogenesis. These mechanisms for longer-term regulation of lipogenesis take several days to become fully manifested and augment the direct and immediate effect of free fatty acids and hormones such as insulin and glucagon.Please help "Biochemistry for Medics" by CLICKING ON THE ADVERTISEMENTS above!