De Novo Synthesis of Fatty Acids

DE NOVO SYNTHESISOF

FATTY ACIDSFloro B. Madarcos, MDDept. of Biochemistry

URMMMC College of Medicine

OBJECTIVES At the end of the lecture, the

student should be able to discuss de novo synthesis of fatty acids in terms of: Definition of the pathway Sites – organs and intracellular Reaction steps and regulation Role of fatty acid multienzyme

complex End product

Modifications and examples Sources of NADPH2

DE NOVO SYNTHESIS OFFATTY ACIDS

I.Definition

The process of combining eight 2-carbonfragments (acetyl groups from acetyl CoA) fo form a 16-carbon saturated

fatty acid palmitate.


II. Cellular Location

Cytoplasm


III. Sites A. Main – liver and adipocytesB. Other Sites

Nervous system Special tissues under specific conditions, such as the mammary glands during lactation

HCO3- + ATP

Citrate lyase

Pyruvate

OxaloacetatePi + ADP

Pyruvatecarboxylase

(biotin)Acetyl-CoA

HS-CoA + NAD

CO2 + NADPH

Pyruvatedehydrogenase

complex

CitrateHS-CoA

Mitochondrial Matrix

H+

CitrateAnion

(Malate, Pyruvate or Pi)

Cytosol

Malate

Oxaloacetate

NAD+

NADH + H+

Malatedehydrogenase

Acetyl CoA

HS-CoA

Pi + ADP

ATP

PyruvateMalic enzyme

NADPH2NADP

IV. REACTIONS OF DE NOVO SYNTHESIS OF FATTY ACIDS

First Step: Production of Cytosolic Acetyl CoA by the Citrate Transport System

Citrate synthase

Pt

To FA Synthesis

CO2

IV. REACTIONS OF DE NOVO SYNTHESIS OF FATTY ACIDS

B. Second Step: Carboxylation of Acetyl CoA to Malonyl CoA

O ||CH3 – C – S – CoA

ACETYL CoA

ATP

ADP + Pi

O O \ || C – CH2 – C – S – CoA //O MALONYL CoA

Acetyl CoA carboxylase(Biotin)

HCO3-

(CO2)

CitrateInsulinHigh CHOLow FatHigh Prot. +

Malonyl CoAPalmitoyl CoAEpinephrineGlucagonHigh FatFasting

- H2O

Mn+2BiotinRegulation1. Short Term Covalent Allosteric2. Long Term

V. FATTY ACID SYNTHASE MULTIENZYME COMPLEX

β-KETO-ACYL

SYNTHASE

ACETYLTRANS ACY-LASE

MALONYLTRANSACYLLASE

HYDRATASE ENOYL

REDUCTASEβ-KETOACYL

REDUCTASE

ACPTHIO

ESTERASE

THIOESTERASE

KETOACYL

REDUCTASE ENOYL REDUCTASE

HYDRATASE

MALONYLTRANSACYLLASE

ACETYLTRANS ACY-LASE

β-KETO-ACYL

SYNTHASE

4-PHOSPHO-PANTETHEINE

SH

CYS

SUBUNITDIVISION

FUNCTIONAL

UNIT

CYS

SH

SH

ACP

SH

4-PHOSPHO-PANTETHEINE

Pantothenic acid

1

2

REACTIONS OF FATTY ACIDSYNTHASE

CYS–SH

ACP–SH

O ||CH3 – C – S – CoA Acetyl CoA

1Acetyl CoA trans-

acylase

CYS–SH O ||ACP–S – C – CH3

Acetyl-S-ACP

0 ||CYS–S-C-CH3 ACP–SH2

0 ||CYS–S-C-CH3

O O || |ACP–S-C-CH2-C |

O O|| ||C-CH2-C-S-CoA|O

Malonyl CoA

3Malonyl trans

acylase Malonyl-S-ACP

CO2

CYS–SH O O || ||ACP–S – C – CH2 – C – CH3

Acetoacetyl – S - ACP

CYS–SH O || H OH ACP–S – C – C – C – CH3

H Hβ- KetohydroxyButyryl –S-ACP

4

5

NADPH+ + H+NADP+

β-KetoacylACP-synthase

β-KetoacylACP-reductase

CYS–SH O || ACP–S – C – CH = CH – CH3

Crotonyl – S - ACP

H2O

β- HydroxylACP-dehydratase

6

NADPH+ + H+

NADP+

7 Enoyl ACP-reductase

ACP– SH

ACP– S-C-CH2

O ||

CYS– S – C - CH2 – CH2 – CH3

O ||

CYS– S – C - CH2 – CH2 – CH3

O O || || C-CH2-C-S-CoA |O

Malonyl CoA

9

O||-C|

O

CYS–SH O || ACP–S – C – CH2 – CH2 – CH3

Butyryl–S-ACP4-Carbon Sat.

Fatty Acyl-ACP CYS–SH O O || ||ACP–S – C – CH2 – C – CH2 – CH2 - CH3

CO2

10

CYS–SH O || ACP–S – C – CH2 – CH2 – CH2 – CH2 - CH3

NADPH+ + H+NADP+NADP+

NADPH+ + H+ H2O

567

CYS–SHACP–S-Palmitate

CYS–SHACP–SHPalmitoyl

thioesteraseRepeat steps

(2) - (7)5x more

6-Carbon sat.Fatty Acyl-ACP 16-Carbon sat.

FA (Palmitoyl ACP) + Palmitate (C16)

VI. REACTIONS OF FATTY ACID SYNTHASE

Butyryl– S-ACP

8

O

CoA1 2

321

12

12 3

124 3

12

12

VI. SYNTHESIS OF PALMITATE (Lippincott’s Biochem, 4TH ed.)

FA synthase

124 36 5

12345

6

Keto group at β-

carbon

α

H2O

STRUCTURE OF PALMITATE (16:0)

O

β α ||CH3–CH2–CH2–CH2–CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-C-O16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

Carboxyl endMethyl end

Acetyl CoA

Malonyl CoA

(16 carbons; no double bond)

VII. SOURCES OF NADPH2 FOR FATTY ACID

SYNTHESISPentose Phosphate Pathway

(HMP)Cytosolic conversion of malate to

pyruvateExtramitochondrial isocitrate

dehydrogenase - ruminants

6 - Phosphogluconate dehydrogenase

NON-OXIDATIVE

STAGE

Glucose 6 – phosphate

6 – phosphogluconate

6 – phosphogluconolactone

Ribulose 5 – phosphate

Ribose 5 – phosphate Xylulose 5 – phosphate

Glyceraldehyde 3 – phosphate

Seduloheptulose 7– phosphate

Fructose 6 – phosphate

Erythrose 4 – phosphate

Fructose 6 – phosphate

Glyceraldehyde 3 – phosphate

GLYCOLYSIS

G6P dehydrogenase

Gluconolactone hydrolase

Transketolase

Transketolase

Transaldolase

OXIDATIVE STAGE

NAPD

NADPH + H

H2O

H+

NAPDNADPH + H

CO2

VIII. PENTOSE PHOSPHATE PATHWAY AS A SOURCE OF NADPH2

HCO3- + ATP

Mitochondrial Matrix

Acetyl CoA

IX. CYTOSOLIC CONVERSION OF MALATE TO PYRUVATE AS A SOURCE OF NADPH2

Citrate lyase

Pyruvate

OxaloacetatePi + ADP

Pyruvatecarboxylase Acetyl-CoA

HS-CoA + NAD

CO2 + NADPH

Pyruvatedehydrogenase

complex

CitrateHS-CoA

H+

CitrateAnion

(Malate, Pyruvate or Pi)

Cytosol

Malate

Oxaloacetate

NAD+

NADH + H+

Malatedehydrogenase

HS-CoA

Pi + ADP

ATP

PyruvateMalate dehydrogenase

(malic enzyme)

NADPH2NADP+

Citrate synthase

Pt

To FA Synthesis

CO2

O-

IC – C – CH2 - IIO

O-

IC IIO

OII

O-

IC – C – CH2 - IIO

O-

IC IIO

OH II

IH

O-

IC – C - IIO

OIIC ICH3

Cytosolic NADH-dependentmalate dehydrogenase

NADH + H+ NAD+

NADP+-dependentmalate dehydrogenase

(malic enzyme)

NADP+

NADPH + H+

CO2

Oxaloacetate

Pyruvate

Malate

CYTOSOLIC CONVERSION OF MALATE TO PYRUVATE AS A SOURCE OF NADPH2

Glyceraldehyde 3-PO4

dehydrogenase reaction(glcolysis)

Reductive synthesis

of FAs, steroids,

sterols

X. BALANCE SHEET FOR THE CONVERSION OF ACETYL CoA

TO PALMITATE

8Acetyl CoA + 7Malonyl CoA + 14NADPH

+ 14H+

1Palmitate + 7CO2 + 8CoA + 14NADP+ + 6H2O

XI. PAMITATE MODIFICATION

Chain Elongation Desaturation Hydroxylation

XII. PAMITATE MODIFICATION

Sites ER – Malonyl CoA/ NADPH/

Palmitoyl CoA Mitochodrion – Acetyl CoA,

NAD & NADPH

XIII. FATTY ACID CHAIN ELONGATION

IN THE ENDOPLASMIC RETICULUMPALMITOYL CoA

β-KETOACID

MALONYL CoA

REDUCTION(similar to de novo

pathway)

SATURATED FA 26 CARBONS (even numbers)

NADPH

XIV. FATTY ACID ELONGATION IN THE MITOCHONDRION

O ||

R – CH2 – C – SCoA (Fatty acyl CoA)

O O | | ||R – CH2 – C – CH2 – C – SCoA

O ||CH3 – C – SCoA

CoASH β-ketothiolase

OH O | ||R – CH2 – CH – CH2 – C – SCoA

β-hydroxyacyl CoAdehydrogenase

NADH + H+

NAD+

O ||R – CH2 – CH = CH – C – SCoA

Enoyl CoAhydratase

H2O

O ||R – CH2 – CH2 – CH2 – C – SCoA

NADP+

NADPH + H+ Enoyl CoAreductase

Acetyl CoA

Fig. 17.12, p. 685, Devlin’s Textbook of Biochemistry, 7th ed.

XV. FATTY ACID DESATURATION

SAT. FATTY ACID

UNSAT. FATTY ACID(monoenoic fatty acid)

Cytochrome b5

NADPH-cytochrome b5 reductase

O2 Δ9

UNSAT. FATTY ACID

UNSAT. FATTY ACID(with more = bonds)

Δ4Δ5Δ6

Fatty acid desaturase

Cytochrome b5

NADPH-cytochrome b5 reductase

O2

XVI. FATTY ACID DESATURATION- EXAMPLES

16:0 (Sat. FA Palmitic acid)

18:0 (Sat. FAStearic acid )

18:2Δ9,12

(Dienoic FA)

16:Δ9 (Unsat. FAPalmitoleic acid)

18:3Δ6,9,12

(Trienoic FA)18:1Δ9 (Unsat. FA

Oleic acid)

Δ9(Stearoyl-CoAdesaturase)

Δ6

ω-7 ω-6ω-9

A B C

Δ9(Stearoyl-CoAdesaturase)

XVII. EXAMPLE OF FATTY ACID DESATURATION

O ||

CH3–CH2–CH2–CH2–CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-C-O16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

O

||

CH3–CH2–CH2–CH2–CH2-CH2-CH = CH- CH2-CH2-CH2-CH2-CH2-CH2-CH2-C-O16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

Palmitate (C16:0); saturatedO2

2 H2O

∆9 desaturase

NADH + H+

NAD+

Palmitoleic acid (16:∆9); monounsaturated; monoenoicomega-7

XVIII. ANOTHER EXAMPLE OF FATTY ACID DESATURATION

O ||

CH3-CH2-CH2- CH2- CH2- CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-C-O18 17 16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

O

||

CH3-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH= CH- CH2-CH2-CH2-CH2-CH2-CH2-CH2-C-O18 17 16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

Stearic acid (C18:0); saturatedO2

2 H2O

∆9 desaturase

NADH + H+

NAD+

Oleic acid (18:∆9); monounsaturated; monoenoicomega-9

XIX. EXAMPLE OF FATTY ACID ELONGATION AND DESATURATION

O

||CH3–CH2- CH2- CH2–CH2–CH = CH-CH2-CH = CH-CH2-CH2-CH2-CH2-CH2-CH2-CH2-C-S-CoA18 17 16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

O2

2 H2O∆6 desaturase

Linoleoyl CoA (18: 2∆9,12); dienoic

NADH+ + H+

NAD+

O

||CH3–CH2- CH2- CH2–CH2–CH = CH-CH2-CH = CH-CH2-CH = CH-CH2-CH2-CH2-CH2-C-S-CoA18 17 16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

γ-Linolenoyl CoA (18: 3∆6,9,12); trienoicMalonyl CoA

CO2 + HS-CoA

Elongase

O

||CH3-CH2-CH2-CH2-CH- CH=CH-CH2-CH=CH2-CH2-CH=CH-CH2-CH2-CH2-CH2-CH2-CH2-C-S-CoA20 19 18 17 16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

Eicosatrienoyl CoA (20: 3∆8,11,14); trienoic

O ||

CH3-CH2-CH2-CH2-CH- CH=CH-CH2-CH=CH-CH2-CH=CH-CH2-CH=CH-CH2-CH2-CH2-C-S-CoA20 19 18 17 16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

Arachidonoyl CoA (20: 4∆5,8,11,14); tetraenoic, omega-6

O2

2 H2O∆5 desaturase

NADH+ + H+

NAD+

Methylene groups

XX. HUMAN DESATURATION OF

FATTY ACIDS

CH3-(CH2)6+n - CH2 - CH2 - CH2 - CH2 - CH2 - CH2 – (CH2)2 -COOH Δ

4 des

atur

ase

Δ5 d

esat

uras

e

Δ6 de

satu

rase

Δ9 de

satu

rase

9

8

7

6

3

2

5

4

1

16

15 - 10

XXI. CAN HUMANS SYNTHESIZE THESE TWO

FATTY ACIDS?

A. CH3 -(CH2)4-CH=CH-CH2-CH=CH-(CH2)7- COOH

B. CH3 -(CH2)4-CH=CH-CH2-CH=CH-CH2-CH=CH-(CH2)4- COOH5

8 1911 1017 13 1218

8 19 7 612 101118 1317

Omega-6

Omega-6

REFERENCES

Horton, Principles of Biochemistry, 4th ed. Devlin, Textbook of Biochemistry, 7th ed. Lippincott’s Biochemistry, 4th ed. Harper’s Illustrated Biochemistry, 28th ed.

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De Novo Synthesis of Fatty Acids