Amino Acids, Proteins, and Enzymes

Lipogenesis &

Lipolysis

Lipogenesis Triacylglycerol synthesis

Definition:

• Lipogenesis is the synthesis of TAG from fatty acids and glycerol.

Site:

• Sub-cellular site: cytoplasm

• Organ (tissue) site: liver (primary site), adipose tissue, lactating mammary gland, kidney

Esters of glycerol with three fatty acids

Functions of Triglyceride

Main stored from of energy in adipose cells. Energy: 9kcal/gram

1) Synthesis of glycerol phosphate (activation of glycerol):

Steps of Lipogenesis

2) Synthesis of acyl-CoA (activation of fatty acids):

Fatty acid ---> acyl CoA

Thiokinase CoA

ATP AMP+Pi

Fates of the formed TAG

In liver: TAG VLDL tissues In adipose tissue: TAG stored as depot

fat

Acyl transferases catalyes the transfer acyl groups

Regulation of lipogenesis

Insulin stimulates lipogenesis

Adipocytes can take glucose only in the presence of insulin

Insulin stimulate glycolysis which supplies glycerol phosphate

Anti-insulin hormones inhibit lipogenesis

is the hydrolysis or breakdown of triacylglycerol in adipose tissue to glycerol and 3 fatty acids

It requires enzymes called lipases which are present in the adipose tissue.

Site:Sub-cellular site: cytoplasm

Lipolysis

The hydrolysis of

triacylglycerol is initiated by hormone sensitive lipase.

HSL is Hormonally regulated

Regulation of lipolysis

• Hormone-sensitive lipase (HSL) is activated when phosphorylated by cyclic AMP-dependent protein kinase

• Epinephrine & other anti-insulin hormones stimulate lipolysis during fasting, stress & hypoglycaemia by 1. activation of adenylate cyclase

forming cAMP

• Insulin inhibits lipolysis by

1. Inhibiting adenylate cyclase enzyme 2. Stimulating phosphodiesterase

3. Stimulating phosphatase enzyme

Lipid Metabolism in Fat Cells:

Fed State Starved State

• Insulin

• Stimulates up-take of glucose by Adipocytes

• stimulates glycolysis

– increased glycerol phosphate synthesis

– increases esterification

• inactivates HSL

• net effect: TG storage

• Glucagon, epinephrine

• activates HSL

• net effect: TG mobilization and increased FFA

• inhibit lipogenesis

FATTY ACIDS SYNTHESIS

Definition: Synthesis of saturated fatty acids Product: palmitate (16 c)

Site:

– Sub-cellular site: cytoplasm

– Organ (tissue) site: liver, lactating mammary gland, adipose tissue & kidney Main requirements:

– Acetyl CoA (Active Acetate) – Acetyl CoA carboxylase enzyme

– ATP for activation & fixation of CO2 in the synthesis of malonyl CoA from acetyl – CoANADPH

ACP Fatty Acid Synthase β-Ketoacyl synthase

β-Ketoacyl synthase Fatty Acid Synthase ACP

S

H SH

SH SH

Fatty acid synthase multienzyme complex.

It is a dimer. Each unit contains 7 enzymes and a protein (acyl carrier protein)

Steps of Fatty acid synthesis

A.

Translocation of acetyl CoA from mitochondria to cytoplasm (Citrate shuttle)

B. Cytoplasmic pathway for fatty acid synthesis:

1.

Carboxylation of acetyl CoA (2 carbon atoms) to malonyl CoA by acetyl CoA carboxylase (ACC)

2.

Formation of palmitate (16 C) by Fatty acid synthase multienzyme complex.

Cytoplasmic pathway for fatty acid synthesis:

• 1- Carboxylation of acetyl CoA by Acetyl CoA Carboxylase

• ‘first reaction’ of fatty acid synthesis

• This is the irreversible regulatory step in fatty acid synthesis

• malonyl-CoA serves as activated donor of acetyl groups in FA synthesis

2- Formation of palmitate (16 C):

All the next steps are catalyzed by Fatty acid synthase

Priming reactions (Transacetylases) (1) Condensation Rxn

(2) Reduction Rxn (3) Dehydration Rxn (4) Reduction Rxn

Thioesterase

The overall reaction for palmitate synthesis:

1 Acetyl CoA+7 Malonyl CoA+14(NADPH+H†)+7ATP---Palmitate(16C)+8CoA+14NADP†+7(ADP+Pi)+7H2O

Pant SH

Cys SH

Pant SH

Cys S C CH₃

O

Pant S

Cys S C CH₃

O C

CH₂ COO^

O

Pant S

Cys SH C

CH₂ C

O

CH₃ O acetyl-S-CoA HS-CoA malonyl-S-CoA HS-CoA CO₂

1 2

1 Malonyl/acetyl-CoA-ACP Transacylase 2 Malonyl/acetyl-CoA-ACP Transacylase

REGULATION OF FATTY ACID SYNTHESIS

Acetyl CoA---> Malonyl CoA

AllostericHormonal

Acetyl CoA Carboxylase

Citrate

Malonyl CoA Palmitoyl CoA

Insuline Glucagon Epinephrine

-

I- short-term regulation of acetyl CoA carboxylase

High-calorie, high- carbohydrate diets Low-calorie diet or fasting

-

+

II- long-term regulation of acetyl CoA carboxylase

+

-

+

Catabolic Pathway Of FA

Beta Oxidation Of Fatty Acids

Beta Oxidation of Fatty acids

•

Beta Oxidation is the process where energy is produced by degradation of fatty acids to acetyl-CoA units

•The process of fatty acid oxidation is termed b oxidation since it occurs through the sequential removal of 2-carbon units (as acetyl-CoA) by oxidation at the b-carbon position of the fatty acyl-CoA molecule.

Importance of b-oxidation 1. Energy production

2. Acetyl-COA production

β -oxidation takes place in Mitochondria

→ Fatty acids which are participating in β-oxidation undergo activation to form Fatty acyl CoA

→ Activation of fatty acid takes place in CYTOPLASM, requires 2 high energy bonds and enzyme is

Thiokinase or fatty acyl Co A synthetase.

1

2. Transport of acyl CoA into the mitochondria ( rate-limiting step) The activated fatty acids which are present in CYTOPLASM

enters into MITOCHONDRIA with the help of CARNITINE CARRIER.

FA~CoA

FA~Carnitine

Acyl transferase I

FA~Carnitine Carnitine

Acyl transferase II

Translocase HS-CoA

FA~CoA HS-CoA

Carnitine

N(CH₃)₃ CH₂

H-C-OH

COO^- CH₂

+

Carnitine

CYTOPLASM

THE MATRIX

3. β – oxidation proper

There are 4 steps in

β C– oxidation

Step I – Oxidation by FAD linked dehydrogenase Step II – Hydration by Hydratase

Step III – Oxidation by NAD linked dehydrogenase Step IV – Thiolytic clevage Thiolase

Beta Oxidation

Beta Oxidation

Beta Oxidation Palmitic (16 c)

21

 -Oxidation and ATP

Activation of a fatty acid requires: 2 ATP

One cycle of oxidation of a fatty acid produces:

• 1 NADH 3 ATP

• 1 FADH₂ 2 ATP

Acetyl CoA entering the citric acid cycle produces:

• 1 Acetyl CoA 12 ATP

Palmitic acid (16 C) needs 7 cycles of beta-oxidation, which gives rise to 8 acetyl Co A x 12 = 96 ATP

7 FADH₂ x 2 = 14 ATP 7 NADH x 3 = 21 ATP

Total 131 ATP

In initial activation Palmic acid → Palmitoyl Co A requires 2 high energy phosphates, so net is 131 – 2 = 129 ATP

• Odd number fatty acids are

oxidized by β-oxidation until the final 3 carbons propionyl CoA is produced.

• So, β- oxidation of odd number fatty acid gives: (n) acetyl CoA + 1 propionyl CoA

• Propionyl CoA is converted to succinyl CoA which can enter the TCA cycle

β- oxidation of odd number fatty acids

Ketone Bodies A special source of fuel and

energy for certain tissues

• Some of the acetyl-CoA produced by fatty acid oxidation in liver mitochondria is converted to acetone, acetoacetate and - hydroxybutyrate

• These are called "ketone bodies"

• Source of fuel for brain, heart and muscle

• Ketogenesis increases in fasting , diabetes and starvation

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Oxidation of Ketone Bodies for Energy

Most tissues can oxidize ketone bodies

-requires mitochondria

Conditions that increase KB synthesis and oxidation

Length of fasting (> 3 days) Low carb diets

Untreated Type 1 diabetes Diabetic ketoacidosis

Liver synthesizes KB but can not oxidize them: liver lacks CoA transferase

ENERGY PRODUCTION FROM OXIDATION OF

-HYDROXYBUTYRATE

Lipid transport

 Non-covalent assemblies of lipids and proteins

 LP core

◦ Triglycerides

◦ Cholesterol esters

 LP surface

◦ Phospholipids

◦ Proteins

◦ cholesterol

Plasma Lipoproteins (Structure)

Function as transport vehicles for triacylglycerols and cholesterol in the blood

All the lipids contained in plasma, including fat, phosphalipids, cholesterol, cholesterol ester and fatty acid, exist and transport in the form of lipoprotein

CM

_{VLDL LDL}

HDL

Lipoprotein Nomenclature, Composition and seperation

Major apoB 48 apoB 100 apoB 100 apoA-I Protein

Major TG TG CE CE Lipid

1. Electrophoresis method:

- Lipoprotein fast pre -Lipoprotein

-Lipoprotein

CM (chylomicron) slow

2. Ultra centrifugation method ：

high density lipoprotein (HDL) high low density lipoprotein ( LDL)

very low density lipoprotein ( VLDL)

CM (chylomicron ) slow

Lipids (Triacylglycerols) are Transported as Lipoproteins

Intestinal Mucosa (Exogenous Lipids) Chylomicrons

Liver (Endogenous Lipids)

VLDL (very low density lipoproteins)

◦Synthesized in small intestine

◦Transport dietary lipids (exogenous TG)

◦98% lipid, large sized, lowest density

◦Apo B-48

 Receptor binding

◦Apo C-II

 Lipoprotein lipase activator

◦Apo E

 Remnant receptor binding

Chylomicrons

Nascent chylomicron (B-48)

Mature chylomicron (+apo C & apo E) Lipoprotein lipase

Chylomicron remnant Apo C removed Removed in liver

◦ Synthesized in liver

◦ Transport endogenous triglycerides

◦ 90% lipid, 10% protein

◦ Apo B-100

 Receptor binding

◦ Apo C-II

 LPL activator

◦ Apo E

 Remnant receptor

binding ^• Nascent VLDL (B-100) + HDL (apo C & E) = VLDL

• LPL hydrolyzes TG forming IDL

• 75% of IDL removed by liver

• 25% of IDL converted to LDL by hepatic lipase

Very Low Density Lipoprotein (VLDL)



Is both consumed (exogenous) and produced by the liver (endogenous).



Endogenous production increases with high ingestion of Saturated fat



Found ONLY in animal products

Blood Cholesterol

Acetyl CoA is the source of all carbon atoms in cholesterol Acetyl CoA is the source of all carbon atoms in cholesterol

Squalene β -hydroxy- β- methylglutaryl

CoA

Mevalonate

Farmesyl pyrophosphate Acetyl CoA

CoA Acetoacetyl CoA

CoA

Acetyl CoA

HMG-CoA reductase

Cyclization

Inhibition



Formation site: from VLDL in blood, but a small part is

directly released from liver



Function: transport cholesterol from liver to the peripheral

tissues.



Carries aprox. 50% of blood cholesterol. containing only apo B-100.



LDL concentration in blood has positive correlation with incidence of cardiovascular diseases.

Low Density Lipoproteins (LDL - Bad)

Fates of cholesterol in the cells

1. Incorporated into cell membranes.

2. Metabolized to steroid hormones.

3. Re-esterified and stored.

4. Expulsion of cholesterol from the cell, and transported by HDL and finally excreted through liver.

 Formation site: liver and intestine

 Function: transport cholesterol from peripheral tissues to liver (reverse cholesterol transport)

 Reservoir of apoproteins

 Contain  protein,  Cholesterol

• Protects against heart disease

◦ Apo A

 Activates lecithin-

cholesterol acyltransferase (LCAT)

◦ Apo C

 Activates LPL

◦ Apo E

 Remnant receptor binding

High Density Lipoproteins (HDL – Good)

1. Uptake of cholesterol from peripheral tissues

2. Esterification of HDL-C by LCAT 3. Transfer of CE to (IDL and CR) 4. removal of CE-rich remnants by

liver

5. converted to bile acids and excreted

a . To combine and transport lipids.

b . To regulate lipoprotein metabolism.

apo A II activates hepatic lipase （ HL ） apo A I activates LCAT

apo C II activates lipoprotein lipase （ LPL ） c. To recognize the lipoprotein receptors.

Functions of apolipoproteins

Apolipoproteins