Fatty Acid Synthesis Fatty Acid Synthase Acetyl-CoA serves as a primer Addition of two-carbon units from malonyl-CoA Each two-carbon unit added must be reduced by 2 NADPH + 2 H+ Reaction for the synthesis of Palmitic acid (C:16): Acetyl-CoA + 7 Malonyl-CoA + 14 NADPH + 14H+ Palmitic acid + 7 CO2 + 14 NADP+ + 8 CoA + 6 H2O

Oxaloacetate + Acetyl-CoA Cytosolic Acetyl-CoA & NADPH Generation (presented as in most text books, this scheme ignores the specificities of mitochondrial transporters; a more accurate description is in the handout) Glycolysis Mitochondrion Acetyl-CoA Pyruvate Citrate Pyruvate Oxaloacetate TCA cycle Malate Citrate Cytosol ATP + CoA Citrate lyase Malate dehydrogenase ADP + Pi Malic enzyme Pyruvate Malate Oxaloacetate + Acetyl-CoA NADPH + H+ + CO2 NADP+ NAD+ NADH+H+ ATP + CO2 Acetyl-CoA carboxylase ADP + Pi Fatty acid synthesis Malonyl-CoA

Fatty Acid Synthesis - O-C-CH2-C-S-CoA Malonyl-CoA is produced by Acetyl-CoA carboxylase Acetyl-CoA (cytoplasmic) + HCO3- Malonyl-CoA O || CH3-C-S-CoA O O || || - O-C-CH2-C-S-CoA ATP ADP + Pi Acetyl-CoA Carboxylase Requires Biotin

Fatty Acid Synthesis Acetyl-CoA Carboxylase Rate limiting reaction for fatty acid synthesis ACC1 is a liver isozyme Small amounts of ACC2 are present in muscle where malonyl-CoA has a regulatory function (Fatty acid oxidation)

Fatty Acid Synthesis Acetyl-CoA Carboxylase 1 Highly regulated Allosteric activation by citrate; inhibition by palmitoyl-CoA. Inhibited by phosphorylation in the fasting state. (low blood glucose inhibits; phosphorylation state is determined by both glucagon activation of a kinase and insulin activation of a phosphatase). Transcriptional up regulation by ChREBP (high carbohydrate diet increases amount of ACC1 and most other enzymes of fatty acid synthetic pathway)

Fatty Acid Synthesis Acetyl-CoA Carboxylase 1 Transcriptional control Xylulose-5-phosphate Acetyl-CoA + Insulin Transcription Citrate Palmitoyl-CoA H2O Pi + ─ Protein phosphatase + CO2 ATP ADP + Pi P P Phosphorylated Acetyl CoA carboxylase PKA AMPK Acetyl CoA carboxylase Acetyl CoA carboxylase + + (Inactive) (Active) ADP + Pi ATP (Inactive) Glucagon AMP Malonyl-CoA Allosteric regulation Covalent modification

Triacylglycerol Synthesis Long-term transcriptional regulation by ChREBP (Carbohydrate Regulatory Element Binding Protein). In addition to short term regulation of Acetyl-CoA carboxylase Many enzymes of fatty acid & triacylglycerol synthetic pathway are coordinately regulated by ChREBP. ChREBP is inhibited by Protein Kinase A dependent phosphorylation. ChREBP is activated by Protein Phosphatase 2A dependent dephophorylation (PP2A is stimulated by Xyulose-5-P). Low Glucose: Glucagon cAMP Protein kinase A Inactive ChREPB-P Fatty acid synthesis High Glucose: Xyulose-5-P Protein Phosphatase A2 Active ChREPB-OH Fatty acid synthesis

Fatty Acid Synthesis The main product of fatty acid synthase is palmitic acid (16:0). Fatty acids can be elongated by other enzymes that add two carbon units from malonyl-CoA. Elongation is particularly important in brain. Still other enzymes can add double bonds (usually at 9 ). Omega-3 and omega-6 fatty acids can not be synthesized by humans.

Triacylglycerol Synthesis Fatty acids must be activated to Acyl-CoA Fatty acid + CoA + ATP Acyl-CoA + AMP + PPi PPi + H2O 2 Pi Acyl-CoA synthetase Pyrophosphatase

Triacylglycerol Synthesis Glycerol-3-phosphate is required for triacylglycerol synthesis. H2C-OH | HOCH O | | H2C-O-P-O - || O - H2C-OH | O=C O | | H2C-O-P-O - || O - Glycerol-3-phosphate dehydrogenase Dihydroxyacetone Phosphate Glycerol-3-phosphate NADH + H+ NAD+ Glycerol-3-phosphate dehydrogenase

Triacylglycerol Synthesis Addition of 3 Acyl groups from Acyl-CoA to Glycerol-3-phosphate Glycerol-3-phosphate Phosphatidate Triacylglycerol 2 Acyl-CoA CoA Acyl-CoA CoA + Pi

VLDL formation Apolipoprotien B-100 has a repeating -helix/-sheet structure: Lipids are packaged as apolipoprotein B-100 is being synthesized: From Shelness & Sellers (2001) Curr Opin Lipidology 12:151-157

VLDL formation VLDL stands for Very Low Density Lipoprotein As it is synthesized, VLDL contains: One molecule of apoliprotein B-100 Triacylglycerol Phospholipid Cholesterol ester Microsomal Triacylglycerol Transfer Protein(MTP) assists in the formation of the VLDL Other components are added to the VLDL in the blood.

VLDL formation Apolipoprotein B-100 synthesis is required for the transport of lipid out of the liver If protein synthesis is reduced (e.g. by malnutrition) fat droplets accumulate in the liver. If the rate of lipid synthesis is greatly elevated with respect to protein synthesis (e.g. in type I diabetes or glucose 6-phosphatase deficiency) fat droplets accumulate in the liver.