BC10-2(part2)

Carboxylation of Acetyl CoA to Form _______

- regulated step in fatty acid synthesis

a. _________

- inactive in multiple carboxylase deficiency (inability to utilize biotin)

i. Requires

- ATP

- HCO3

-

- coenzymes _______ (covalently bound to the lysyl residue of the carboxylase)

1. ___________-Term Regulation of Acetyl CoA Carboxylase

- this carboxylation reaction is both

- rate-limiting

- regulated step in fatty acid synthesis

a. ____________ Regulation

- inactive form of acetyl CoA carboxylase (consists of protomer made of 4 subunits) 

allosteric activation by citrate  polymerization of protomers

- enzyme is inactivated by (___________ - the end product of the pathway)

- __________ (intermediate in the pathway)

- __________ (end-product of the pathway)

 depolymerization of the protomer

b. Reversible Phosphorylation

i. __________, __________

 enzyme phosphorylation  enzyme inactivation

ii. __________

 acetyl CoA carboxylase dephosphorylation  enzyme activation

iii. ______________

 enzyme phosphorylation

- allosterically activated by a rise in AMP relative to ATP

- covalently activated by phosphorylation via AMPK kinase

2. Long-Term Regulation of Acetyl CoA Carboxylase

a. High ……… Diet (High _________-High ________ Diets)

 increased enzyme synthesis  increased fatty acid synthesis

b. _______ Calorie Diet, _______

 decreased enzyme synthesis  decreased fatty acid synthesis (fatty acid synthase is also regulated by this mode of dietary manipulation)

D. Fatty Acid Synthase (Palmitate Synthase): a Multienzyme Complex

1. Eukaryotes

- consist of a dimer (each monomer has 7 different enzymatic activities + a domain that covalently binds a molecule of 4’-phosphopantetheine)

a. 4’-Phosphopantetheine

- derivative of pantothenic acid

- component of acyl carrier protein (ACP)

- carries acetyl and acyl units on its terminal thiol (-SH) group during fatty acid synthesis

D. Fatty Acid Synthase (Palmitate Synthase): a Multienzyme Complex

2. Reactions

a. Transfer of Acetate

- from acetyl CoA to the -SH group of ACP

- catalyzed by acetyl CoA-ACP acetyltransacylase

Acetyl CoA + ACP  Acetyl-S-ACP + CoA

b. Transfer of the Two-Carbon Fragment

- to a cysteine residue on the enzyme (temporary holding site)

Acetyl-S-ACP + Enzyme  Acetyl-S-Enzyme + ACP

c. Transfer of Malonate

- vacant ACP accepts a 3 carbon malonate unit from malonyl CoA

- catalyzed by malonyl CoA-ACP transacylase

Malonyl CoA + ACP  Malonyl-S-ACP + CoA-SH

d. One-Carbon Loss from Malonyl Group

- malonyl group loses the HCO3

-

originally added by acetyl CoA carboxylase 

nucleophilic attack on the thioester bond linking the acetyl group to the cysteine

residue  result in a 4-carbon unit attached to the ACP domain

- loss of free energy from decarboxylation  drives the reaction

- catalyzed by 3-ketoacyl-ACP synthase

Malonyl-S-ACP + Acetyl-S-ACP  Acetoacetyl-S-ACP + ACP + CO2

D. Fatty Acid Synthase (Palmitate Synthase): a Multienzyme Complex

*Next 3 reactions convert the 3-ketoacyl group to the corresponding saturated acyl group by a pair of reductions requiring NADPH and a dehydration steps

e. Alcohol Formation

- keto group is converted to an alcohol

- catalyzed by 3-ketoacyl-ACP reductase

- NADPH is required

Acetoacetyl-S-ACP + NADPH + H+ 

3-Hydroxybutyryl-ACP + NADP+

f. Dehydration Reaction

- water molecule is removed to introduce a double bond

- catalyzed by 3-hydroxyacyl-ACP dehydratase

3-Hydroxybutyryl-S-ACP  Crotonyl-S-ACP + H2O

g. Second Reduction

- catalyzed by enoyl-ACP reductase

- NADPH is required

Crotonyl-S-ACP + NADPH + H+

 Butyryl-S-ACP + NADP+

D. Fatty Acid Synthase (Palmitate Synthase): a Multienzyme Complex

3. Result

- production of a 4 carbon compound (fully saturated)

4. Elongation

- butyryl-ACP reacts with another malonyl group via the reactions described above  cycle

continues

- repetition of the 7 steps incorporating 2 carbon unit (repeated 7x)  16-carbon chain (still

attached to ACP-SH)  process terminated  fully saturated molecule of palmitate

5. Release of Palmitate (16:0)

- catalyzed by palmitoyl thioesterase (deacylase)

- cleaves thioester bond

Palmitoyl-S-ACP + H2O  Palmitate + ACP-SH

6. Overall Reaction for the Synthesis of Palmitate

8Acetyl CoA + 14NADPH + 14H+

+ 7ATP 

Palmitic Acid + 8CoA-SH + 14NADP+

+ 7ADP + 7Pi + 7H2O

- all of the carbons of palmitic acid have passed through malonyl CoA except the 2 donated by the original acetyl CoA (found at the methyl group-end of the fatty

acid)

E. Major Sources of the NADPH Required for Fatty Acid Synthesis

1. ____________ Pathway

- major supplier of NADPH

- ___ NADPH produced per glucose molecule entering this pathway

2. ____________

-Dependent Malate Dehydrogenase (Malic Enzyme)

- oxidize and decarboxylate malate  pyruvate

- source of cytosolic NADPH

- malate can arise from the reduction of OAA by cytosolic NADP+

-dependent malate dehydrogenase

- cytosolic NADH can be produced during glycolysis

F. Interrelationship Between Glucose Metabolism and Palmitate Synthesis

1. _________

- produces

- pyruvate (primary source of mitochondrial acetyl CoA to be used for fatty acid synthesis)

- cytosolic reducing equivalents of NADH

2._________

- produced in the 1st step in the gluconeogenic pathway

3. _________

- produced in the mitochondria

- condenses with OAA  citrate (1st step in the TCA cycle)

4. _________

- leaves the mitochondria

- cleaved in the cytosol to produce cytosolic acetyl CoA

5. _________

- produced during glycolysis

- contribute to the reduction of NADP+

to NADPH needed for palmitoyl CoA synthesis

6. _________ of Cytosolic Acetyl CoA

- used to synthesize _________(NADPH as the source of reducing equivalents for the pathway)