Voet D., Voet Ju.G. Biochemistry

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step and therefore “costs” the free energy of GTP hydroly-

sis. The liver lacks 3-ketoacyl-CoA transferase, which per-

mits it to supply ketone bodies to other tissues.

4 FATTY ACID BIOSYNTHESIS

Fatty acid biosynthesis occurs through condensation of C

units, the reverse of the ␤-oxidation process. Through iso-

topic labeling techniques, David Rittenberg and Konrad

Bloch demonstrated, in 1945, that these condensation units

are derived from acetic acid.Acetyl-CoA was soon proven

to be a precursor of the condensation reaction, but its

mechanism remained obscure until the late 1950s when

Salih Wakil discovered a requirement for bicarbonate in

fatty acid biosynthesis and malonyl-CoA was shown to be

an intermediate. In this section, we discuss the reactions of

fatty acid biosynthesis.

A. Pathway Overview

The pathway of fatty acid synthesis differs from that of

fatty acid oxidation. This situation, as we saw in Section

18-1D, is typically the case of opposing biosynthetic and

degradative pathways because it permits them both to be

thermodynamically favorable and independently regulated

under similar physiological conditions. Figure 25-29 out-

lines fatty acid oxidation and synthesis with emphasis on

the differences between these pathways.Whereas fatty acid

oxidation occurs in the mitochondrion and utilizes fatty

acyl-CoA esters, fatty acid biosynthesis occurs in the

cytosol with, as Roy Vagelos discovered, the growing fatty

acids esterified to acyl-carrier protein (ACP; Fig. 25-30).

ACP, like CoA, contains a phosphopantetheine group that

forms thioesters with acyl groups.The phosphopantetheine

phosphoryl group is esterified to a Ser OH group of ACP,

whereas in CoA it is esterified to AMP. In animals, ACP is

part of a large multifunctional protein (Type I ACP; see

below), whereas in E. coli it is a 125-residue polypeptide

(Type II ACP). The phosphopantetheine group is trans-

ferred from CoA to apo-ACP to form the active holo-ACP

by phosphopantetheine transferase (alternatively, ACP

synthase).

The redox coenzymes of the animal fatty acid oxidative

and biosynthetic pathways differ (NAD



and FAD for ox-

idation; NADPH for biosynthesis) as does the stereochem-

istry of their intermediate steps, but their main difference is

the manner in which C

units are removed from or added

to the fatty acyl thioester chain. In the oxidative pathway,

-ketothiolase catalyzes the cleavage of the C



¬C



bond

of -ketoacyl-CoA so as to produce acetyl-CoA and a new

fatty acyl-CoA, which is shorter by a C

unit. The G°¿ of

this reaction is very close to zero so it can also function in

the reverse direction (ketone body formation). In the

biosynthetic pathway, the condensation reaction is coupled

to the hydrolysis of ATP, thereby driving the reaction to

completion. This process involves two steps: (1) the ATP-

dependent carboxylation of acetyl-CoA by acetyl-CoA

carboxylase (ACC) to form malonyl-CoA, and (2) the

Section 25-4. Fatty Acid Biosynthesis 961

. Indeed, the breath of individuals with ketosis (also

called ketoacidosis), a potentially pathological condition in

which acetoacetate is produced faster than it can be metab-

olized (a symptom of diabetes; Section 27-4B), has the

characteristic sweet smell of acetone.

The liver releases acetoacetate and -hydroxybutyrate,

which are carried by the bloodstream to the peripheral tis-

sues for use as alternative fuels. There, these products are

converted to acetyl-CoA as is diagrammed in Fig. 25-27.

The proposed reaction mechanism of 3-ketoacyl-CoA

transferase (Fig. 25-28), which catalyzes this pathway’s sec-

ond step, involves the participation of an active site car-

boxyl group both in an enzyme–CoA thioester intermedi-

ate and in an unstable anhydride. Succinyl-CoA, which acts

as the CoA donor in this reaction, can also be converted to

succinate with the coupled synthesis of GTP in the succinyl-

CoA synthetase reaction of the citric acid cycle (Section

21-3Ea). The “activation” of acetoacetate bypasses this

Figure 25-28 Proposed mechanism of 3-ketoacyl-CoA

transferase involving an enzyme–CoA thioester intermediate.

C E

–

C E

–

SCo

–

SCoA

Succinyl-CoA

–

O C

Unstable anhydride

Enzyme–CoA

thioester intermediate

•

–

SCoACH

OCH

Unstable anhydride

•

–

CCO

–

Succinate

ECoAS C

Acetoacetate

Acetoacetyl-CoA

JWCL281_c25_940-1018.qxd 4/20/10 1:59 PM Page 961

962 Chapter 25. Lipid Metabolism

the degradation of leucine; Section 26-3F). The ACC reac-

tion, like those of the other biotin-dependent carboxylases,

occurs in two steps, a CO

activation and a carboxylation:

E biotin

–

C CH

SCoA

Biotinyl–enzyme

Carboxybiotinyl–enzyme

Malonyl-CoA

Acetyl-CoA

HCO

–

+ ATP

ADP

+ P

E biotin CO

–

exergonic decarboxylation of the malonyl group in the

condensation reaction catalyzed by fatty acid synthase.

These enzymes are discussed below.

B. Acetyl-CoA Carboxylase

ACC is a biotin-dependent enzyme that catalyzes the first

committed step of fatty acid biosynthesis and one of its rate-

controlling steps. It is a member of a family of biotin-

dependent carboxylases that, in humans, has only three

members besides ACC: propionyl-CoA carboxylase

(Section 25-2Ea), pyruvate carboxylase (Fig. 23-4), and

-methylcrotonyl-CoA carboxylase (which participates in

Figure 25-29 Comparison of fatty acid  oxidation and fatty

acid biosynthesis. Differences occur in (1) cellular location,

(2) acyl group carrier, (3) electron acceptor/donor, (4) stereo-

Figure 25-30 The phosphopantetheine group in acyl-carrier protein (ACP) and in CoA.

FADH

NAD

NADH + H

CoA

Acetyl-CoA

CoA + CO

Malonyl-CoA

NADP

NADPH + H

NADP

NADPH + H

FAD

Fatty acyl-CoA (C

n +2

)

Fatty acyl-ACP (C

n +2

)

Fatty acyl-CoA (C

)

Fatty acyl-ACP (C

)

Enoyl-CoA

Enoyl-ACP

L-Hydroxyacyl-CoA

D-Hydroxyacyl-ACP

β-Ketoacyl-CoA

β-Ketoacyl-ACP

Biosynthesis

Oxidation

Occurs in mitochondrion Occurs in cytoplasm

CoA is acyl

group carrier

ACP is acyl

group carrier

FAD is electron

acceptor

NADPH is

electron donor

NAD

is electron

acceptor

NADPH is

electron donor

unit product

is acetyl-CoA

unit donor

is malonyl-CoA

L-β-Hydroxyacyl

group

D-β-Hydroxyacyl

group

chemistry of the hydration/dehydration reaction, and (5) the

form in which C

units are produced/donated. See the

Animated Figures

Cysteamine

Phosphopantetheine prosthetic group of ACP

N C

N CH

ACPO SerO

OH CH

–

Cysteamine

Phosphopantetheine group of CoA

N C

N CH

PCH

O OO

OH CH

–

Adenine

2–

PO OH

JWCL281_c25_940-1018.qxd 4/20/10 1:59 PM Page 962

E. coli ACC is a multienzyme complex in which these

steps are catalyzed by separate subunits: the homodimer

biotin carboxylase (BC; 456 residues) and the 



heterotetramer carboxyltransferase (CT; 319 and 304

residues). In addition, the biotin is linked as a biocytin

residue (Fig. 23-3b) to the biotin carboxyl-carrier protein

(BCCP; 156 residues), which forms homodimers. In con-

trast, mammalian and avian ACCs contain both enzy-

matic activities as well as the biotin carboxyl carrier on a

single 2346-residue polypeptide chain in the order

BC–BCCP–CT (which differs from the order in pyruvate

carboxylase, which is BC–CT–BCCP; Section 23-1Ae).

The structure of an intact ACC has not been determined,

although the X-ray structure the E. coli BC subunit

closely resembles the BC domain of pyruvate carboxylase

(Fig. 23-5a). Interestingly, however,the CT domains of the

various biotin-dependent carboxylases differ greatly in

sequence and structure.

a. Acetyl-CoA Carboxylase Is Regulated by

Hormonally Controlled Reversible Phosphorylation

ACC is subject to hormonal regulation. Glucagon as well

as epinephrine and norepinephrine (adrenaline and nora-

drenaline; Section 18-3Ea) trigger the enzyme’s cAMP-

dependent increase in phosphorylation, which inactivates

the enzyme. Insulin, on the other hand, stimulates enzyme

dephosphorylation and thus its activation.

The mechanism by which cAMP causes an increase in

the phosphorylation state of ACC is interesting. ACC is

phosphorylated, in vitro, by two different kinases, the

cAMP-dependent protein kinase A (PKA; Section 18-

3Cb) at Ser 77 and AMP-dependent protein kinase

(AMPK; Sections 25-5a and 27-1) (which is cAMP inde-

pendent) at Ser 79, 1200, and 1215.Yet, when liver cells are

incubated with cAMP-elevating hormones in the presence

P-ATP, Ser 77 is found not to be labeled. Evidently, an

increase in [cAMP] results in a phosphorylation increase

at sites modified by AMPK rather than by PKA. How can

this be? It appears that, in vivo, the cAMP-dependent in-

crease in phosphorylation occurs not through the phos-

phorylation of new sites but, rather, through the inhibition

of dephosphorylation of previously phosphorylated posi-

tions. We have already seen such a mechanism in operation

in the control of glycogen metabolism,where the cAMP-de-

pendent phosphorylation of phosphoprotein phosphatase

inhibitor-1 causes the inhibition of dephosphorylation

(Section 18-3C). In the case of ACC, however, dephospho-

rylation is catalyzed by phosphoprotein phosphatase-2A,

which is not affected by phosphoprotein phosphatase in-

hibitor-1. The mechanism by which PKA causes the in-

crease in phosphorylation associated with AMPK activity

is, as yet, unknown.

b. Avian and Mammalian Acetyl-CoA Carboxylases

Undergo Enzyme Polymerization on Activation

Electron microscopy reveals that both avian and mam-

malian ACCs form long filaments of 20 to 40 protomers

(Fig. 25-31). This polymeric form of the enzyme is catalyti-

cally active but the protomer is not. The rate of fatty acid

biosynthesis is therefore controlled by the position of the

equilibrium between these forms:

Phosphorylation favors the inactive protomer while dephos-

phorylation favors the active polymer. Several metabolites

also affect the activity of acetyl-CoA carboxylase. Citrate

promotes the polymerization of ACC, whereas palmitoyl-

CoA and other fatty acyl-CoA’s promote its depolymeriza-

tion. Thus, cytosolic citrate, whose concentration increases

when the mitochondrial acetyl-CoA concentration builds up

(Section 25-4D), activates fatty acid biosynthesis and hence

is a feedforward activator, whereas palmitoyl-CoA, the

pathway product, is a feedback inhibitor.

c. Mammalian Acetyl-CoA Carboxylase

Has Two Major Isoforms

There are two major isoforms of mammalian ACC.ACC1

occurs in adipose tissue and ACC2 occurs in tissues that

oxidize but do not synthesize fatty acids, such as heart muscle.

Tissues that both synthesize and oxidize fatty acids, such as

liver,contain both isoforms, which are homologous although

the genes encoding them are located on different chromo-

somes. What is the function of ACC2? The product of the

ACC-catalyzed reaction, malonyl-CoA, strongly inhibits

Protomer(inactive) Δ polymer(active)

Section 25-4. Fatty Acid Biosynthesis 963

Figure 25-31 Association of acetyl-CoA carboxylase

protomers. An electron micrograph with an accompanying

interpretive drawing indicates that filaments of avian liver

acetyl-CoA carboxylase consist of linear chains of flat rectangular

protomers. [Courtesy of Malcolm Lane,The Johns Hopkins

University School of Medicine.]

400Å

JWCL281_c25_940-1018.qxd 6/8/10 8:59 AM Page 963

SCoA

+ H SACP

Acetyl-CoA

SACP

Acetyl-ACP

SCoA

SACPH

H+

SACP

Acetoacetyl-ACP

+ NADPH

NADP

SACP

OOH

D-

-Hydroxybutyryl-ACP

α,β

-trans-Butenoyl-ACP

+ NADPH

NADP

SACP

Butyryl-ACP

SACP

Palmitoyl-ACP

(CH

)

SACP

Palmitate

–

palmitoyl thioesterase (TE)

enoyl-ACP reductase (ER)

β-ketoacyl-ACP reductase (KR)

β-ketoacyl-ACP synthase (KS)

malonyl/acetyl-CoA-ACP

transacylase (MAT)

SCoA

+ H SACP

Malonyl-CoA

SACP

Malonyl-ACP

malonyl/acetyl-CoA-ACP

transacylase (MAT)

–

SACP

β-hydroxyacyl-ACP dehydrase (DH)

SCoA

(CH

)

after 7 reaction cycles

recycle Reactions 2a–5

six more times

964 Chapter 25. Lipid Metabolism

the mitochondrial import of fatty acyl-CoA for fatty acid

oxidation, the major control point for this process. Thus it

appears that ACC2 has a regulatory function (Section 25-5).

Prokaryotic acetyl-CoA carboxylases are not subject to

any of these controls.This is because fatty acids in these or-

ganisms are not stored as fats but function largely as phos-

pholipid precursors.The E. coli enzyme is instead regulated

by guanine nucleotides so that fatty acids are synthesized

in response to the cell’s growth requirements.

C. Fatty Acid Synthase

The synthesis of fatty acids from acetyl-CoA and malonyl-

CoA involves seven enzymatic reactions that yield mainly

palmitic acid (see below). These reactions were first studied

in cell-free extracts of E. coli, in which they are catalyzed by

independent enzymes together with ACP.These proteins are

collectively known as type II fatty acid synthase (FAS-II).

FAS-II also occurs in chloroplasts, the only site of fatty acid

synthesis in plants, and in vertebrate mitochondria, whose

components are all nuclear encoded and closely resemble

their bacterial counterparts. In fungi,however,fatty acids are

synthesized by a 2600-kD ␣

␤

multifunctional enzyme,

whereas in animals they are synthesized by a multifunctional

enzyme consisting of two identical ⬃275-kD polypeptide

chains, each containing all seven activities plus ACP and

known as type I fatty acid synthase (FAS-I). Most of the do-

mains catalyzing these reactions in these so-called megasyn-

thases are homologous to the corresponding proteins that

constitute FAS-II, which indicates that they arose through

the joining of what were previously independent proteins.

The reactions catalyzed by FAS-I to synthesize palmi-

tate are diagrammed in Fig. 25-32. The long flexible phos-

phopantetheine chain of ACP (Fig. 25-30) functions to

transport the substrate between the protein’s various enzy-

matic domains:

1a. The transfer of the acetyl group from acetyl-CoA to

ACP to yield acetyl-ACP as catalyzed by malonyl/acetyl-

CoA-ACP transacylase (MAT).

Figure 25-32 Reaction cycle for the biosynthesis

of fatty acids. The biosynthesis of palmitate requires

seven cycles of C

elongation followed by a final

hydrolysis step.

See the Animated Figures

JWCL281_c25_940-1018.qxd 8/9/10 9:43 AM Page 964

2a. The loading of -ketoacyl-ACP synthase (KS; also

known as condensing enzyme) by the transfer of the acetyl

group from ACP to a KS Cys residue, thus maintaining the

acetyl group’s thioester linkage.

1b. The formation of malonyl-ACP in a reaction analo-

gous to that of Reaction 1a, which in animals is catalyzed

by the same enzyme, MAT.

2b. The coupling of the acetyl group to the C



of the

malonyl group on the ACP with the malonyl group’s ac-

companying decarboxylation so as to form acetoacetyl-

ACP and free the KS active site Cys-SH group (Fig. 25-33).

Consequently, the CO

taken up in the acetyl-CoA carboxy-

lase reaction (Section 25-4B) does not appear in the product

fatty acid. Rather, the decarboxylation functions to drive

carbon–carbon bond formation in the condensation reac-

tion, which through the acetyl-CoA carboxylase reaction, is

coupled to ATP hydrolysis.

3–5. The reduction, dehydration, and further reduc-

tion of acetoacetyl-ACP so as to form butyryl-ACP as se-

quentially catalyzed by -ketoacyl-ACP reductase (KR),

-hydroxyacyl-ACP dehydrase (DH), and enoyl-ACP re-

ductase (ER). The coenzyme in both reductive steps is

NADPH, whereas in  oxidation, the analogs of Reactions

3 and 5, respectively, use NAD



and FAD (Fig. 25-29).

Moreover, Reaction 3 produces and Reaction 4 requires a

D--hydroxyacyl group, whereas the analogous reactions in

 oxidation involve the corresponding

L isomer.

2a to 5 Repeat. The butyryl group from the butyryl-

ACP is transferred to the Cys-SH of KS. Thus the acetyl

group with which the system was initially loaded has been

elongated by a C

unit. The ACP is “reloaded” with a mal-

onyl group (Step 1b), and another cycle of C

elongation

occurs. This process occurs altogether seven times to yield

palmitoyl-ACP.

6. The palmitoyl-ACP thioester bond is hydrolyzed by

palmitoyl thioesterase (TE), yielding palmitate, the normal

product of the fatty acid synthase pathway, and regenerat-

ing the enzyme for a new round of synthesis.

The stoichiometry of palmitate synthesis therefore is

Since the 7 malonyl-CoA are derived from acetyl-CoA as

follows:

the overall stoichiometry for palmitate biosynthesis is

a. Animal FAS-I Is a Flexible X-Shaped Dimer

Most but not all of the enzymatic activities of animal

FAS-I remain functional when this dimeric enzyme is dis-

sociated into monomers. Moreover, fragments resulting

from the limited proteolysis of animal FAS-I exhibit many

of the enzymatic activities of the intact protein. Appar-

ently, contiguous stretches of its polypeptide chain fold to

form a series of autonomous domains, each with a specific

but different catalytic activity. Several other enzymes, for

example, mammalian acetyl-CoA carboxylase (Section

25-4B), exhibit similar multifunctionality but none has as

many separate catalytic activities as does animal FAS-I. The

order of the domains along the animal FAS-I polypeptide

chain is indicated in Fig. 25-34. Three of these domains

besides ACP lack enzymatic activity: a linker domain

Palmitate  14NADP



 8CoA  6H

O  7ADP  7P

8 Acetyl-CoA  14NADPH  7ATP

7 malonyl-CoA  7ADP  7P

 7H



7 Acetyl-CoA  7CO

 7ATP

Palmitate  7CO

 14NADP



 8CoA  6H

Acetyl-CoA  7 malonyl-CoA  14NADPH  7H



Section 25-4. Fatty Acid Biosynthesis 965

S ACP

Malonyl-ACP

Acetoacetyl-CoA

–

S Cys KS

HS Cys KS

–

S ACP

Figure 25-33 The mechanism of carbon–carbon bond forma-

tion in fatty acid biosynthesis. The condensation of an acetyl

group on the active site Cys of -ketoacyl-ACP synthase (KS)

with a malonyl group on the phosphopantetheine arm of ACP

forms a -ketoacyl-ACP.The reaction is driven by the exergonic

elimination of CO

from the malonyl group to generate a reso-

nance-stabilized acetyl-ACP carbanion intermediate that func-

tions as a good nucleophile.

Figure 25-34 Domain organization of porcine FAS-I at approximate sequence scale. [Modified

from a drawing by Timm Maier and Nenad Ban, ETH Zurich, Switzerland.]

JWCL281_c25_940-1018.qxd 6/8/10 8:59 AM Page 965

966 Chapter 25. Lipid Metabolism

(LD) that bridges the KS and MAT domains; a pseudo-

methyltransferase (ME) domain, so named because it is

homologous to the methyltransferase family; and a pseudo-

ketoreductase (KR) domain, so named because it is a

truncated form of the KR domain.

The X-ray structure of porcine FAS-I in complex with

NADP



, determined by Nenad Ban, reveals a pseudo-2-

fold symmetric, X-shaped dimer (Fig. 25-35). Its two sub-

units associate via an extensive interface involving over

150 residues per chain from the KS, ER, and DH domains.

However the upper and lower portions of the X are only

loosely connected.The lower portion of the X contains the

enzyme’s two condensing activities, MAT and KS (which

catalyze reactions 1 and 2 in Fig. 25-32), whereas the upper

portion of the X contains the enzyme’s -carbon modifying

activities, DH, KR,and ER (which catalyze reactions 3–5 in

Fig. 25-32). The C-terminal ACP and TE domains are flexi-

bly tethered to the enzyme and are therefore unobserved.

The KR–ACP linker consists of ⬃13 residues and can

therefore span a distance of up to ⬃40 Å, whereas the

ACP–TE linker has a length of ⬃25 residues and can there-

fore extend ⬃80 Å.

Each of the two reaction chambers of porcine FAS-I is

lined by the full set of catalytic domains from one sub-

unit, all of which must be visited by the phosphopan-

tethiene arm of an ACP to carry out all the FAS reac-

Figure 25-35 X-ray structure of porcine FAS-I in complex

with NADP



. (a) The ⬃190-Å wide homodimer, as viewed

perpendicular to its pseudo-2-fold axis (vertical black arrow),

with its various domains colored as in Fig. 25-34 and its linkers

gray. The bound NADP



cofactors are drawn in space-filling

form in blue. The attachment sites of the disordered ACP/TE

(a)

(b)

domains are indicated by black dots.The domain names of the

second subunit have appended primes. (b) A corresponding

schematic diagram indicating how the various domains are

linked. [Courtesy of Timm Maier and Nenad Ban, ETH Zurich,

Switzerland. PDBid 2VZ9.]

JWCL281_c25_940-1018.qxd 6/8/10 8:59 AM Page 966

tions. The catalytic sites of these domains occur on both

the front and back faces of the protein and their distribu-

tion has no discernable relationship to the order of the

reactions in the catalytic cycle. In the structure shown in

Fig. 25-35a, the length of the KR–ACP linker would al-

low the ACP to reach all the catalytic sites in the same re-

action chamber but not those in the opposite reaction

chamber (e.g., the distance between the ACP attachment

site on one subunit and the MAT domain in the other is

⬃135 Å). Nevertheless, mutational studies by Stuart

Smith in which ACP on one subunit and MAT or KS on

the other subunit were inactivated yielded functional en-

zymes albeit with reduced activities, thus indicating that

an ACP domain can service both MAT domains and both

KS domains. The most plausible explanation for these

observations is that the upper portion of the X can rotate

180° with respect to the lower portion. This hypothesis is

supported by the observation that the quite tenuous

DH/KS contact that joins the top and bottom portions of

the X lacks perfect 2-fold symmetry in Fig. 25-35a, thus

indicating that this joint is flexible.

b. Fungal FAS-I Has a Barrel-Like Shape

The reactions catalyzed by 



fungal FAS-I differ

from those mediated by animal FAS-I in several respects:

1. The bifunctional MAT activity of animal FAS-I

transfers both acetyl and malonyl groups from CoA to

ACP (Reactions 1a and 1b of Fig. 25-32). However, fungal

FAS-I employs a monofunctional acetyl transferase (AT)

activity to transfer the incoming acetyl group from CoA to

ACP and a malonyl/palmitoyl transferase (MPT) activity

to do so for malonyl groups.

2. Animal FAS-I synthesizes an ACP-linked palmitoyl

group, which its TE activity hydrolytically releases as

palmitate (Reaction 6 of Fig. 25-32). In contrast, fungal

FAS-I synthesizes ACP-linked palmitoyl (C

) and stearoyl

) groups in a ratio of roughly 2:3, which its bifunctional

MPT activity then transfers to CoA to yield the pathway’s

final products, palmitoyl-CoA and stearoyl-CoA.

3. The ER activity in animal FAS-I uses NADPH to

directly reduce the C“C double bond in Reaction 5 of

Fig. 25-32. However, in fungal ER, NAPDH reduces FMN

to FMNH

, which in turn, reduces the double bond.

4. Fungal FAS-I has a phosphopantetheine transferase

(PPT) activity that attaches the phosphopantetheine group

to ACP.This activity is not part of animal FAS-I.

The distribution of the various enzymatic activities along

the  and  chains of fungal FAS-I (Fig. 25-36) bears no re-

semblance to that of animal FAS-I (Fig. 25-34). Note that

the MPT activity is shared by both the  and  chains.

The X-ray structure of FAS-I from the thermophilic

fungus Thermomyces lanuginosus (Fig. 25-37), determined

by Ban, reveals that it forms a hollow barrel-shaped protein

with D

symmetry (the rotational symmetry of a trigonal

prism; Fig. 8-65b). The six  chains form a D

-symmetric

wheel that is capped on each side by a 3-fold symmetric

dome that is predominantly formed by the  chains. The

central wheel splits the hollow interior of the barrel into

two identical reaction chambers, each of which has several

openings in its side walls through which small molecules

can enter. The active sites of all the enzymatic activities

face the interiors of the reaction chambers. Clearly, animal

and fungal FAS-I’s have divergently evolved from FAS-II.

Section 25-4. Fatty Acid Biosynthesis 967

Figure 25-36 Domain organization of fungal FAS-I at approximate sequence scale. [Modified

from a drawing by Simon Jenni and Nanad Ban, ETH Zurich, Switzerland.]

Figure 25-37 X-ray structure of T. lanuginosus FAS. The

⬃270-Å-high by ⬃250-Å-diameter 



heterododecamer is

viewed along a 2-fold axis (ellipsoid) and perpendicular to its

3-fold axis (triangle) with its various domains colored as in Fig.

25-36 and its linker domains gray. The attachment sites of the

disordered ACP domains are indicated by black dots. [Courtesy

of Simon Jenni and Nenad Ban, ETH Zurich, Switzerland.

PDBids 2UV9 and 2UVA.]

JWCL281_c25_940-1018.qxd 6/8/10 8:59 AM Page 967

968 Chapter 25. Lipid Metabolism

The six ACP domains, which are N-terminally anchored

to the chamber walls and C-terminally anchored to the

middle of the central wheel, are disordered in the X-ray

structure of T. lanuginosus FAS-I. However, they are visi-

ble in the otherwise closely similar X-ray structure of yeast

FAS-I, which was independently determined by Ban and

by Thomas Steitz. Structural considerations suggest that

each doubly tethered ACP domain can swing to visit the re-

quired six catalytic centers, which in the case of the ACP

tethered to subchamber 1 are KR from 1, KS from 2,

MPT and DH from 1, and AT and ER from 2, where

subchamber 2 is on the clockwise side of subchamber 1 as

viewed from the top of the dome.

The PPT domains are located on the outside of the bar-

rel where they cannot interact with the ACP domains. This

suggests that they attach the phosphopantetheine groups

to the ACPs before the barrel has fully assembled.

c. Fatty Acid Synthase Inhibitors

Are Drug Candidates

In well-nourished individuals, fatty acid synthesis pro-

ceeds at a low rate. However, certain tissues, particularly

cancers, express high levels of FAS-I and produce fatty

acids at a high rate. Consequently, inhibitors of animal

FAS-I are being investigated as possible anticancer agents.

Moreover,the differences between the enzymatic activities

of the various types of FAS’s, particularly their ER activi-

ties, makes FAS-II and fungal FAS-I targets for the devel-

opment of novel antibiotics.

d. Variations on a Theme: Polyketide Biosynthesis

Polyketides are a family of 10,000 diverse and struc-

turally complex natural products, many of which have anti-

bacterial, antifungal, antitumor, and immunosuppressive

properties, that are synthesized by bacteria, fungi, plants,

and certain marine animals. They are made by the modular

condensation of acyl-CoA monomers such as acetyl-CoA

and propionyl-CoA with malonyl-CoA and methylmalonyl-

CoA extender units whose decarboxylation drives the

condensation reaction. The name polyketide comes from

the fact that the primary condensation products have

-keto functional groups. Palmitate is an example of a

polyketide since it is formed by the condensation of one

acetyl-CoA primer and seven malonyl-CoA extender

units. Following each condensation reaction, the new

-keto group may be reduced, dehydrated, and reduced

again as with fatty acids, or may undergo only partial

modification.

Polyketides are synthesized by megasynthases. We

have already seen that animal FAS-I contains seven enzy-

matic activities as well as ACP. Another example of a

polyketide is 6-deoxyerythronolide B (6dEB), the parent

macrolactone of the antibiotic erythromycin A (Section

32-3G), which is synthesized in the soil bacterium Saccha-

ropolyspora erythraea from one propionyl-CoA primer

and six (S)-methylmalonyl-CoA extenders by deoxyery-

thronolide B synthase (DEBS; Fig. 25-38). DEBS is a

2000-kD, 





complex of 3000-residue subunits

whose three homodimeric units each catalyze two elonga-

tion/modification cycles. Unlike FAS-I, which catalyzes

several cycles of elongation/modification with the same

active sites, DEBS catalyzes each elongation/modification

cycle on a different module, which permits the differences

in the modifications that occur at each cycle. Thus, DEBS,

which has 28 different active sites, functions much like an

assembly line. Module 4, as Fig. 25-38 indicates, is almost

identical in function to FAS-I, containing KS, AT, ACP,

KR, DH, and ER, and reducing its primary -ketone con-

densation product to a methylene group. However, it does

not contain TE because the elongation process is not

complete after this phase. Module 3 contains only ACP,

KS, and AT, the minimal set of activities for a module, and

passes its -ketone condensation product to module 4

without further modification. Modules 1, 2, 5, and 6 con-

tain only ACP,AT, KS, and KR, the sites necessary for the

condensation and ketone reduction steps, thereby gener-

ating hydroxy products. The overall organization of the

modules therefore creates a polyhydroxy product con-

taining one keto group and one methylene group in the

chain. The DEBS final product, 6dEB, is a lactone pro-

duced by the reaction of the terminal hydroxyl group with

the thioester anchoring the growing chain to the synthase.

The various polyketide synthases have different organiza-

tions of modules, and consequently synthesize a multi-

tude of different compounds.

D. Transport of Mitochondrial Acetyl-CoA

Into the Cytosol

Acetyl-CoA is generated in the mitochondrion by the ox-

idative decarboxylation of pyruvate as catalyzed by pyru-

vate dehydrogenase (Section 21-2A) as well as by the oxi-

dation of fatty acids. When the need for ATP synthesis is

low, so that the oxidation of acetyl-CoA via the citric acid

cycle and oxidative phosphorylation is minimal, this mito-

chondrial acetyl-CoA may be stored for future use as fat.

Fatty acid biosynthesis occurs in the cytosol but the mito-

chondrial membrane is essentially impermeable to acetyl-

CoA. Acetyl-CoA enters the cytosol in the form of citrate via

the tricarboxylate transport system (Fig. 25-39). Cytosolic

ATP-citrate lyase then catalyzes the reaction

which resembles the reverse of the citrate synthase reac-

tion (Section 21-3A) except that ATP hydrolysis is

required to drive the intermediate synthesis of the “high-

energy” citryl-CoA, whose hydrolysis drives the citrate

synthase reaction to completion. ATP hydrolysis is there-

fore required in the ATP-citrate lyase reaction to power

the resynthesis of this thioester bond. Oxaloacetate is re-

duced to malate by malate dehydrogenase. Malate may be

oxidatively decarboxylated to pyruvate by malic enzyme

(Section 25-2Ed) and be returned in this form to the mi-

tochondrion. The malic enzyme reaction resembles that

acetyl-CoA  oxaloacetate  ADP  P

Citrate  CoA  ATP Δ

JWCL281_c25_940-1018.qxd 4/20/10 1:59 PM Page 968

of isocitrate dehydrogenase in which a ␤-hydroxy acid is

oxidized to a ␤-keto acid, whose decarboxylation is

strongly favored (Section 21-3C). Malic enzyme’s coen-

zyme is NADP

⫹

, so when this route is used NADPH is

produced for use in the reductive reactions of fatty acid

biosynthesis.

Citrate transport out of the mitochondrion must be bal-

anced by anion transport into the mitochondrion. Malate,

pyruvate, and P

can act in this capacity. Malate may there-

fore also be transported directly back to the mitochon-

drion without generating NADPH.As we have seen in Sec-

tion 25-4C, synthesis of each palmitate ion requires 8

molecules of acetyl-CoA and 14 molecules of NADPH. As

many as 8 of these NADPH molecules may be supplied

with the 8 molecules of acetyl-CoA if all the malate pro-

duced in the cytosol is oxidatively decarboxylated. The re-

maining NADPH is provided through the pentose phos-

phate pathway (Section 23-4).

E. Elongases and Desaturases

Palmitate (16:0), the normal product of the animal fatty acid

synthase pathway, is the precursor of longer chain saturated

and unsaturated fatty acids through the actions of elongases

and desaturases. Elongases are present in both the mito-

chondrion and the ER but the mechanisms of elongation at

the two sites differ.Mitochondrial elongation (a process in-

dependent of the fatty acid synthase pathway) occurs by

successive addition and reduction of acetyl units in a rever-

sal of fatty acid oxidation; the only chemical difference be-

tween these two pathways occurs in the final reduction step

in which NADPH takes the place of FADH

as the terminal

Section 25-4. Fatty Acid Biosynthesis 969

Figure 25-38 An example of polyketide biosynthesis: the synthesis of erythromycin A. [After

Pfeifer, B.A., Admiraal, S.J., Gramajo, H., Cane, D.E., and Khosla, C., Science 291, 1790 (2001).]

Erythromycin A

OH OH

SSS S SS

N(CH

)

DEBS: Deoxyerythronolide B synthase

KS: Ketosynthase

T: Acyltransferase

CP: Acyl carrier protein

KR: Keto reductase

ER: Enoyl reductase

DH: Dehydratase

TE: Thioesterase

Elongation 1

Elongation 2

Elongation 3

Elongation 4

Elongation 5

Elongation 6

glycosylation/

hydroxylation

OCH

6-Deoxyerythronolide B

(6dEB)

Propionyl-CoA

(S)-Methylmalonyl-CoA

release/cyclization

AT ACP KS AT KR ACP KS AT KR ACP KS AT ACP KS AT DH ER KR

KS AT KR ACP ACPKS AT KR TE

ACP

Module 1 Module 3

DEBS1 (370 kD) DEBS2 (380 kD) DEBS3 (332 kD)

Module 5 Release

Module 2 Module 4 Module 6

SCoA

COOH

SCoA

JWCL281_c25_940-1018.qxd 8/9/10 9:44 AM Page 969

970 Chapter 25. Lipid Metabolism

redox coenzyme (Fig. 25-40). Elongation in the ER in-

volves the successive condensations of malonyl-CoA with

acyl-CoA. These reactions are each followed by NADPH-

associated reductions similar to those catalyzed by FAS-I,

the only difference being that the fatty acid is elongated as

its CoA derivative rather than as its ACP derivative.

Unsaturated fatty acids are produced by terminal de-

saturases. Mammalian systems contain four terminal

desaturases of broad chain-length specificities designated



-, 

-, and 

-fatty acyl-CoA desaturases. These

Figure 25-39 Transfer of acetyl-CoA from mitochondrion to

cytosol via the tricarboxylate transport system.

Figure 25-40 Mitochondrial fatty acid elongation. Elongation occurs by the

reversal of fatty acid oxidation with the exception that the final reaction

employs NADPH rather than FADH

as its redox coenzyme.

Mitochondrion Cytosol

Inner

mitochondrial

membrane

NAD

ATP

ATP-citrate lyase

ADP

malate dehydrogenase

NADH + H

+ CO

malic enzyme

NADP

Acetyl CoA

ADP

citrate synthase

pyruvate carboxylase

ATP

HCO

–

C O

–

HHO

–

Pyruvate

Malate

Oxaloacetate

Citrate

CHO CO

–

SCoA

NADPH

SCoACCH

CHO CO

–

Citrate

Pyruvate

C O

–

Oxaloacetate

C O

–

C O

–

SCoACCH

Tricarboxylate

transport

system

SCoA

H SCoA

Acyl-CoA (C

)

Acyl-CoA (C

n+2

)

SCoA

Acetyl-CoA

thiolase

enoyl-CoA hydratase

enoyl-CoA reductase

L-hydroxyacyl-CoA

dehydrogenase

-Ketoacyl-CoA

+ NADH

NAD

+ NADPH

NADP

L--Hydroxyacyl-CoA

SCoA

,-trans-Enoyl-CoA

RCH

SCoA

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