Biosynthesis of Membrane Lipids

Membrane Lipids: Glycerolipids Glycerophospholipid

Glycerophospholipid Head Groups

Membrane Lipids: Sphingolipids Sphingomyelins with phosphocholine or phosphoethanolamine Neutral Glycolipids cerebrosides (1 sugar) globosides (> 2 sugars) Gangliosides complex carbohydrates with sialic acid (Neu5Ac)

Biosynthesis of Membrane Lipids Glycerolipids Sphingolipids Cholesterol

Biosynthesis of Phosphatidic Acid ATP ADP glycerol kinase NADH NAD+ glycerol 3-phosphate dehydrogenase

Biosynthesis of Glycerophospholipids Strategy 1: Prokaryotes: all glycerophospholipids Eukaryotes: phosphatidylinositol phosphatidyglycerol cardiolipin phosphatidylserine (yeast) Strategy 2: phosphatidylcholine phosphatidylethanolamine

Glycerophospholipid Biosynthesis in E. coli Strategy 1 (CDP-DAG) phosphatidylserine (PS) by serine replacing CMP phosphatidylethanolamine (PE) by decarboxylation of PS phosphatidylcholine (PC) by methylation (3x) of PE phosphatidylglycerol (PG) by glycerol 3-phosphate replacing CMP, then phosphatase cardiolipin by one PG replacing glycerol on other PG

Biosynthesis in Eukaryotes of Anionic Glycerophospholipids Strategy 1 (CDP-DAG) phosphatidylglycerol (PG) by glycerol 3-phosphate replacing CMP, then phosphatase cardiolipin by PG replacing CMP on CDP-DAG [CDP-DAG instead of PG] phosphatidylinositol (PI) by inositol replacing CMP phosphorylation of PI at positions 4 and 5

Cardiolipin Biosynthesis Summary Phosphatidylglycerol Glycerol CDP-diacylglycerol cardiolipin synthase (prokaryotic) cardiolipin synthase (eukaryotic) CMP

Biosynthesis of Phosphatidylcholine and Phosphatidylethanolamine in Mammals Strategy 2: CDP-alcohol choline is phosphorylated and cytidylated to form CDP-choline phosphatidylcholine (PC) formed by diacylglycerol replacing CMP phosphatidylethanolamine (PE) formed by analogous pathway starting with ethanolamine salvage pathways for choline and ethanolamine in yeast

Biosynthesis of Phosphatidylserine in Mammals Head group exchange Mammals cannot directly make phosphatidylserine (PS) PS formed by exchanging serine for ethanolamine on PE (endoplasmic reticulum) Mammals can decarboxylate PS to form PE (mitochondria) PC can be made from PE in mammalian liver Salvage pathways in yeast

Summary of Pathways to Phosphatidylcholine and Phosphatidyethanolamine Enzymes for PE and PC: kinases cytidylate transferases DAG transferases methyltransferases (in liver) Also in Mammals: PE ↔ PS exchange PS → PE decarboxylation Not in Mammals: direct PS biosynthesis from CDP-DAG + serine

Biosynthesis of Glycerophospholipids Summary of Strategies: CDP-diacylglycerol + alcohol (head group) CDP-alcohol + diacylglycerol Head group exchange Head group modification (methylation, decarboxylation)

Sphingolipid Biosynthesis Serine decarboxylated and condensed on acyl-CoA NADPH reduces resulting ketone Mixed-function oxygenase forms double bond of sphingosine UDP-glucose for cerebroside PC exchange for sphingomyelin

Cholesterol Biosynthesis Cholesterol is made in 4 stages: Condensation of Mevalonate from 3 Acetates Conversion of Mevalonate into Two Activated Isoprenes Polymerization of 6 Activated Isoprenes into Squalene Cyclization of Squalene and Modification of Lanosterol

Cholesterol Biosynthesis Stage 1: Condensation of Mevalonate from Acetate Final step in β-oxidation of fatty acids in reverse (cytosolic) Aldol condensation at C3 carbonyl to form HMG-CoA Reduction of HMG-CoA Committed step in biosynthesis of isoprenes Requires 2 NADPH for reduction of carboxylate to alcohol

Cholesterol Biosynthesis Stage 2: Conversion of Mevalonate to Activated Isoprenes Requires 3 ATP’s in 4 enzymatic steps

Cholesterol Biosynthesis Stage 3: Polymerization of Activated Isoprenes Farnesyl-PP requires: 1 Dimethylallyl-PP 2 Δ3-Isopentenyl-PP (head to tail polymerization) Squalene requires: 2 farnesyl-PP (head to head polymerization) 1 NADPH required

Cholesterol Biosynthesis Stage 4: Cyclization of Squalene and Modification of Lanosterol Monooxygenase forms squalene 2,3-epoxide Cyclase reaction: H+ opens epoxide ring Cascade of 4 carbocation additions to C=C’s form the 4 rings 2 hydride migrations, 2 methyl migrations, and H+ loss gives lanosterol Modification of lanosterol (19 steps) gives cholesterol

Cholesterol Biosynthesis This Slide FYI only – Not on Final Exam Cholesterol Biosynthesis Stage 4: Conversion of Lanosterol to Cholesterol 19-Step process involves: Oxidative removal of 3 methyl groups as HCO2H or CO2 10 Monooxygenase reactions Oxidation of 15 NAD(P)H Reduction of 2 NAD+ Overall Cholesterol Biosynthesis: 18 ATP hydrolyzed 27 NAD(P)H oxidized (net) from Risley 2002, J. Chem. Educ. 79: 377

Metabolic Fates of Cholesterol 7α-hydroxylase and desmolase are cytochrome P-450 monooxygenases cholesterol 7-dehydrocholesterol 7-dehydrocholesterol reductase 7α-hydroxylase desmolase hν OH cholecalciferol (Vitamin D3) 7α-hydroxycholesterol pregnenolone Bile (Salts) Acids Catabolism Steroid Hormones Vitamin D

Cytochrome P-450 Monooxygenases usually located in smooth endoplasmic reticulum involved in hydroxylation of steroids or xenobiotics General Reaction: AH + BH2 + O–O → A–OH + B + H2O

Biosynthesis of Pregnenolone Steroid hormone synthesis from cholesterol side chain removed in mitochondria of steroidogenic tissues Desmolase is a cytochrome P-450 mixed-function oxidase (monooxygenase) 2 O2 introduce diols at C20, C22 3rd oxidation cleaves the C–C bond with ketone and aldehyde products

Steroid Hormones Pregnenolone

Vitamin D Metabolism in skin: • 7-dehydrocholesterol absorbs ultraviolet B (~300 nm) • previtamin D3 isomerizes to cholecaliferol (vitamin D3) in liver: • vitamin D3 → 1-hydroxyvitamin D3 [1-(OH)D3] in kidney: • 1-(OH)D3 → 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] Final 2 steps involve cytochrome P-450 monooxygenases

Bile (Salts) Acids 7 hydroxycholesterol hydroxylated and oxidized carboxylate is activated with CoA amino groups of glycine or taurine attack activated carboxylate trihydroxycoprostanoate 7α-hydroxycholesterol glycine taurine cholyl CoA glycocholate taurocholate OH