Biosynthesis of Membrane Lipids, Cholesterol, Steroids and Isoprenoids

CH353 February 5, 2008

Summary

• Review of membrane lipid structures and nomenclature– Lehninger Chapter 10

• Biosynthesis of membrane lipid components– Lehninger Chapters 21.3, 21.4

• Membrane Lipids– glycerolipids– sphingolipids– sterols (cholesterol)

• Other Complex Lipids– steroids– isoprenoids

Membrane Lipids: Glycerolipids

Glycerophospholipid

Plasmalogen (ether lipid)

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)

Membrane Lipids: Sterols

• Sterols are polymerized from isoprene units

• Rigid 4-ring structure• Membrane sterols:

– Cholesterol (animals)– Stigmasterol (plants)– Ergosterol (fungi)– None in bacteria

Biosynthesis of Membrane Lipids

• Glycerolipids• Sphingolipids• Cholesterol

Biosynthesis of Phosphatidic Acid

ATPADP

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

CMP

cardiolipin synthase(prokaryotic)

cardiolipin synthase(eukaryotic)

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)

Biosynthesis of Ether Lipids & Plasmalogens

• 2 NADPH required for reducing carboxylate to alcohol

• 1 NADPH for reducing dihydroxyacetone phosphate

Biosynthesis of Ether Lipids and Plasmalogens

• CDP-Ethanolamine substrate for CDP-ethanolamine transferase (correction)

• Long chain alcohol in ether linkage oxidized with mixed-function oxidase (monooxygenase) CDP-ethanolamine

transferase

CDP-ethanolamine

CDP

• 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

Sphingolipid Biosynthesis

Sphingolipid Biosynthesis

Ceramide

Ceramide

UDP- Glucose

UDP

UDP- Galactose

UDP

Glc Gal

Ceramide

UDP- N-Acetylgalactoseamine

UDP

Glc Gal Gal

Ceramide Glc Gal Gal

NAc

Neu

NAcCMP- Sialic Acid

CMP

GM2, a ganglioside

Cholesterol Biosynthesis

Cholesterol is made in 4 stages:

1. Condensation of Mevalonate from 3 Acetates

2. Conversion of Mevalonate into Two Activated Isoprenes

3. Polymerization of 6 Activated Isoprenes into Squalene

4. Cyclization of Squalene and Modification of Lanosterol

Cholesterol Biosynthesis

Stage 1: Condensation of Mevalonate from Acetate

1. Final step in β-oxidation of fatty acids in reverse (cytosolic)

2. Aldol condensation at C3 carbonyl to form HMG-CoA

3. 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

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

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

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

This Slide FYI only – Not on Final Exam

Metabolic Fates of Cholesterol

OH

7α-hydroxycholesterol

cholesterol

pregnenolone

7α-hydroxylase desmolase

7-dehydrocholesterol

7-dehydrocholesterolreductase

hν

cholecalciferol(Vitamin D3)

Bile (Salts) AcidsCatabolism

Steroid Hormones Vitamin D

7α-hydroxylase and desmolase are cytochrome P-450 monooxygenases

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

trihydroxycoprostanoate7α-hydroxycholesterol

glycine taurine

cholyl CoA

glycocholate taurocholate

OH

Isoprenoid Compounds and Derivatives

Isoprenoid Biosynthesis

Dimethylallyl-PPΔ3-Isopentenyl-PP

Geranyl-PP

Farnesyl-PP

Geranylgeranyl-PP

Oligoprenyl-PP

SqualeneStigmasterolLanosterolErgosterol

Cholesterol

Carotenoids

x (3 – 7)

Polyprenyl-PP

x (8 – 21)

Phytyl-PPChlorophyllTocopherols (Vitamin E)Phylloquinone (Vitamin K)

PlastoquinoneUbiquinone (Coenzyme Q)

Dolichol

x 2

Retinoids (Vitamin A)x 2

Mevalonate

Vitamin DBile Salts

Steroid Hormones

HMG-CoA

Statins

C10

C15

C20

C30-50

C55-120

Inhibitors of HMG-CoA Reductase

• Statins: synthetic analogs of mevalonate• Competitive inhibitors of HMG-CoA reductase• For inhibiting cholesterol synthesis

Study Problem

• Statins are widely prescribed drugs for lowering high cholesterol which may lead to atherosclerosis

• They are effective in preventing synthesis of cholesterol by inhibiting HMG-CoA reductase

• Since statins effectively block the entire isoprenoid pathway, there are concerns of potential side effects

• What possible metabolic consequences may statins have by inhibiting isoprenoid biosynthesis?

• What dietary supplements may be prescribed for overcoming possible side effects of statins?