Fatty Acid Synthesis in Eukaryotes

Acetyl-CoA Transport and Fatty Acid Synthesis

• In non-photosynthetic eukaryotes, acetyl-CoA is made in the mitochondria and transported into the cytosol for fatty acid synthesis at a cost of 2 ATPs.
• The total cost of fatty acid synthesis is 3 ATPs per 2C unit, which is approximately 50% of the NADPH+H+ units necessary for fatty acid biosynthesis.

Overview of Fatty Acid Synthesis

• Fatty acids are synthesized in several passes, with each pass involving the reduction of a carbonyl carbon to a methylene carbon.
• The acetate unit for fatty acid synthesis comes from activated malonate in the form of malonyl-CoA.

The Acetyl-CoA Carboxylase Reaction

• The acetyl-CoA carboxylase reaction is the committed step in fatty acid synthesis.

Fatty Acid Synthase (FAS)

• FAS I is a single polypeptide chain with 7 active sites in vertebrates, leading to a single product: palmitate (16:0).
• FAS II is a separate enzyme system in plants and bacteria, producing a variety of fatty acid products (saturated, unsaturated, branched, etc.).

Synthesis of Fat (Triacylglycerides) and Phospholipids

• Animals and plants store fat (triacylglycerides) for fuel.
• The typical 70-kg human has approximately 15 kg of fat, which can last for 12 weeks.
• Animals, plants, and bacteria make phospholipids for cell membranes.
• Both molecules contain a glycerol backbone and 2 (phospholipids) or 3 (triacylglycerides) fatty acids.

Synthesis of Backbone of TAGs and Phospholipids

• Most glycerol 3-phosphate comes from dihydroxyacetone phosphate (DHAP) via glycerol 3-phosphate dehydrogenase.
• Some glycerol 3-phosphate is made from glycerol via glycerol kinase (minor pathway in liver and kidney, absent in adipocytes).

Catabolism and Anabolism of Fatty Acids

• Catabolism of fatty acids produces acetyl-CoA and reducing power (NADH+H+, FADH2) in the mitochondria.
• Anabolism of fatty acids requires acetyl-CoA and malonyl-CoA, as well as reducing power from NADPH+H+, and occurs in the cytosol in animals and chloroplasts in plants.

Fatty Acid Biosynthesis - Place of Action

• Fatty acid synthesis takes place in the cytosol of every cell with mitochondria, although the rate differs between cells.

Fatty Acid Synthesis and NADPH+H+ Levels

• Fatty acid synthesis occurs in cell compartments where NADPH+H+ levels are high, such as in the cytosol of animals and yeast.
• Sources of NADPH+H+ include the pentose phosphate pathway and malic enzyme in adipocytes, and the pentose phosphate pathway in hepatocytes and mammary gland.

The Triacylglycerol Cycle

• 75% of free fatty acids (FFAs) released by lipolysis are reesterified to form TAGs, rather than being used for fuel under all metabolic conditions.
• Some recycling occurs in adipose tissue, where FFAs from adipose cells are transported to the liver, remade into TAG, and redeposited in adipose cells.

Glyceroneogenesis

• Glyceroneogenesis makes DHAP for glycerol 3-phosphate generation during the TAG cycle.
• It is an abbreviated version of gluconeogenesis in the liver and adipose tissue, converting pyruvate to DHAP.
• It uses pyruvate, alanine, glutamine, or any substances from the Citric acid cycle as precursors for glycerol 3-phosphate.

Fatty Acid Biosynthesis

• Animals and plants store fat for fuel, with a typical 70-kg human having ~15 kg fat, enough to last 12 weeks.
• Both TAGs and phospholipids contain a glycerol backbone and 2 or 3 fatty acids, respectively.

Synthesis of Tag and Phospholipids

• Most glycerol 3-phosphate comes from siphoning off DHAP from glycolysis via glycerol 3-phosphate dehydrogenase.
• Some glycerol 3-phosphate is made from glycerol via glycerol kinase, a minor pathway in the liver and kidney, but absent in adipocytes.

Synthesis of Phosphatidic Acid

• Phosphatidic acid is the precursor for TAGs and phospholipids, with fatty acids attached by acyl transferases after activation by acyl-CoA synthetase.
• Phosphatidic acid can be made into triacylglycerol or phospholipid.

Phosphatidic Acid Modifications

• Phosphatidic acid phosphatase (lipin) removes the 3-phosphate from phosphatidic acid, yielding 1,2-diacylglycerol.
• The third carbon is then acylated with a third fatty acid, yielding triacylglycerol.

Regulation of Triacylglycerol Synthesis

• Insulin stimulates triacylglycerol synthesis.
• Lack of insulin results in increased lipolysis, fatty acid oxidation, and sometimes ketone production.
• Failure to synthesize fatty acids from carbohydrates and proteins leads to increased rates of fat oxidation and weight loss in untreated diabetes.

The Triacylglycerol Cycle

• 75% of free fatty acids (FFAs) released by lipolysis are reesterified to form TAGs, rather than being used for fuel under all metabolic conditions.
• Some recycling occurs in adipose tissue, where FFAs from adipose cells are transported to the liver, remade into TAG, and redeposited in adipose cells.

Glyceroneogenesis

• Glyceroneogenesis makes DHAP for glycerol 3-phosphate generation during the TAG cycle.
• It is an abbreviated version of gluconeogenesis in the liver and adipose tissue, converting pyruvate to DHAP.
• It uses pyruvate, alanine, glutamine, or any substances from the Citric acid cycle as precursors for glycerol 3-phosphate.

Fatty Acid Biosynthesis

• Animals and plants store fat for fuel, with a typical 70-kg human having ~15 kg fat, enough to last 12 weeks.
• Both TAGs and phospholipids contain a glycerol backbone and 2 or 3 fatty acids, respectively.

Synthesis of Tag and Phospholipids

• Most glycerol 3-phosphate comes from siphoning off DHAP from glycolysis via glycerol 3-phosphate dehydrogenase.
• Some glycerol 3-phosphate is made from glycerol via glycerol kinase, a minor pathway in the liver and kidney, but absent in adipocytes.

Synthesis of Phosphatidic Acid

• Phosphatidic acid is the precursor for TAGs and phospholipids, with fatty acids attached by acyl transferases after activation by acyl-CoA synthetase.
• Phosphatidic acid can be made into triacylglycerol or phospholipid.

Phosphatidic Acid Modifications

• Phosphatidic acid phosphatase (lipin) removes the 3-phosphate from phosphatidic acid, yielding 1,2-diacylglycerol.
• The third carbon is then acylated with a third fatty acid, yielding triacylglycerol.

Regulation of Triacylglycerol Synthesis

• Insulin stimulates triacylglycerol synthesis.
• Lack of insulin results in increased lipolysis, fatty acid oxidation, and sometimes ketone production.
• Failure to synthesize fatty acids from carbohydrates and proteins leads to increased rates of fat oxidation and weight loss in untreated diabetes.

Acetyl-CoA Transport and Fatty Acid Synthesis

• In non-photosynthetic eukaryotes, acetyl-CoA is made in the mitochondria and transported into the cytosol for fatty acid synthesis at a cost of 2 ATPs.
• The total cost of fatty acid synthesis is 3 ATPs per 2C unit, which is approximately 50% of the NADPH+H+ units necessary for fatty acid biosynthesis.

Overview of Fatty Acid Synthesis

• Fatty acids are synthesized in several passes, with each pass involving the reduction of a carbonyl carbon to a methylene carbon.
• The acetate unit for fatty acid synthesis comes from activated malonate in the form of malonyl-CoA.

The Acetyl-CoA Carboxylase Reaction

• The acetyl-CoA carboxylase reaction is the committed step in fatty acid synthesis.

Fatty Acid Synthase (FAS)

• FAS I is a single polypeptide chain with 7 active sites in vertebrates, leading to a single product: palmitate (16:0).
• FAS II is a separate enzyme system in plants and bacteria, producing a variety of fatty acid products (saturated, unsaturated, branched, etc.).

Synthesis of Fat (Triacylglycerides) and Phospholipids

• Animals and plants store fat (triacylglycerides) for fuel.
• The typical 70-kg human has approximately 15 kg of fat, which can last for 12 weeks.
• Animals, plants, and bacteria make phospholipids for cell membranes.
• Both molecules contain a glycerol backbone and 2 (phospholipids) or 3 (triacylglycerides) fatty acids.

Synthesis of Backbone of TAGs and Phospholipids

• Most glycerol 3-phosphate comes from dihydroxyacetone phosphate (DHAP) via glycerol 3-phosphate dehydrogenase.
• Some glycerol 3-phosphate is made from glycerol via glycerol kinase (minor pathway in liver and kidney, absent in adipocytes).

Catabolism and Anabolism of Fatty Acids

• Catabolism of fatty acids produces acetyl-CoA and reducing power (NADH+H+, FADH2) in the mitochondria.
• Anabolism of fatty acids requires acetyl-CoA and malonyl-CoA, as well as reducing power from NADPH+H+, and occurs in the cytosol in animals and chloroplasts in plants.

Fatty Acid Biosynthesis - Place of Action

• Fatty acid synthesis takes place in the cytosol of every cell with mitochondria, although the rate differs between cells.

Fatty Acid Synthesis and NADPH+H+ Levels

• Fatty acid synthesis occurs in cell compartments where NADPH+H+ levels are high, such as in the cytosol of animals and yeast.
• Sources of NADPH+H+ include the pentose phosphate pathway and malic enzyme in adipocytes, and the pentose phosphate pathway in hepatocytes and mammary gland.

Fatty Acid Synthesis

• The overall goal is to attach an acetate unit (2C) from malonyl-CoA to a growing chain and then reduce it
• The reaction involves cycles of four enzyme-catalyzed steps:
  • Condensation of the growing chain with activated acetate
  • Reduction of carbonyl to hydroxyl
  • Dehydration of alcohol to trans-alkene
  • Reduction of alkene to alkane
• The growing chain is initially attached to the enzyme via a thioester linkage

Acyl Carrier Protein (ACP)

• ACP serves as a shuttle in fatty acid synthesis
• Contains a covalently attached prosthetic group, 4'-phosphopantetheine
• Has a flexible arm to tether the acyl chain while carrying intermediates from one enzyme subunit to the next
• Delivers acetate (in the first step) or malonate (in all the next steps) to the fatty acid synthase

Charging

• Involves the addition of Acetyl-CoA to ACP and further transfer to –SH of KS in synthesis denovo
• Also involves the addition of Malonyl-CoA to ACP
• By using activated malonyl groups in the synthesis of fatty acids and activated acetate in their degradation, the cell makes both processes energetically favorable, although one is effectively the reversal of the other

Enzymatic Activities

• β-ketoacyl-ACP synthase (KS): coupling condensation to decarboxylation of malonyl-CoA makes the reaction energetically favorable
• β-ketoacyl-ACP reductase (KR): reduces β-ketoacyl-ACP to form D-β-hydroxyacyl-ACP
• β-hydroxyacyl-ACP dehydratase (DH): dehydrates D-β-hydroxyacyl-ACP to form trans-Δ2-enoyl-ACP
• Enoyl-ACP reductase (ER): reduces trans-Δ2-enoyl-ACP to form acyl-ACP

Synthesis of Phosphatidic Acid and Triacylglycerol

• Phosphatidic acid is the precursor for TAGs and phospholipids
• Fatty acids are attached by acyl transferases, after activation by acyl-CoA synthetase
• Phosphatidic acid can be modified to form phospholipids or TAGs
• Phosphatidic acid phosphatase (lipin) removes the 3-phosphate from phosphatidic acid to yield 1,2-diacylglycerol

Regulation of Triacylglycerol Synthesis by Insulin

• Insulin stimulates triacylglycerol synthesis
• Lack of insulin results in increased lipolysis, increased fatty acid oxidation, and failure to synthesize fatty acids from carbohydrates and proteins

Triacylglycerol Cycle

• 75% of free fatty acids released by lipolysis are reesterified to form TAGs
• Some recycling occurs in adipose tissue, and some FFAs are transported to the liver, remade into TAG, and redeposited in adipose cells

Glyceroneogenesis

• Generates DHAP for glycerol 3-phosphate generation during TAG cycle
• Occurs during lipolysis, when glycolysis in adipocytes is inhibited
• Converts pyruvate to DHAP using pyruvate, alanine, glutamine, or any substances from the Citric acid cycle as precursors for glycerol 3-phosphate