Structure of heme.

Hemes C and N
atoms are derived
from those of glycine
and acetate.
The mechanism of action of
the PLP-dependent enzyme
-aminolevulinate synthase.
(1) transimination, (2) PLP-
stabilized carbanion formation,
(3) CC bond formation,
(4) CoA elimination,
(5) decarboxylation
facilitated by the PLPSchiff
base, (6) transimination
yielding ALA and regenerating
the PLPenzyme.
A possible mechanism for porphobilinogen synthase.
(1) Schiff base formation, (2) second Schiff base formation,
(3) formation of a carbanion to a Schiff base, (4) cyclization
by an aldol-type condensation, (5) elimination of the enzyme
NH2 group, (6) tautomerization.
The synthesis of uroporphyrinogen III from
PBG as catalyzed by porphobilinogen deaminase
and uroporphyrinogen III synthase. (1a) General
base-catalyzed elimination of NH3 to form a
methylene pyrrolinene intermediate. (1b) Addition to
the methylene pyrrolinene intermediate of the
enzymes covalently linked dipyrromethane cofactor
to form a covalent adduct. (24) Sequential addition
of a second, third, and fourth PBG through successive
NH3 eliminations from PBG to form methylene
pyrrolinene, as in Reaction 1a, followed by addition of
a pyrrole ring carbon atom from the growing chain, as
in Reaction 1b.
(5) Hydrolysis of the methylbilaneenzyme to yield hydroxymethylbilane and
regenerate the free enzymedipyrromethane complex.

(7) Spontaneous cyclization of

hydroxymethylbilane in the
absence of uroporphyrinogen
III synthase. A and P represent
acetyl and propionyl groups.

(6) Synthesis of uroporphyrinogen III via a spiro intermediate by

porphobilinogen deaminase and uroporphyrinogen III synthase.
 Porphobilinogen Deaminase Has a Dipyrromethane
Cofactor

Porphobilinogen deaminase contains a unique dipyrromethane cofactor

(two pyrroles linked by a methylene bridge; rings C1 and C2), which is
covalently linked to the enzyme via a CS bond to an enzyme Cys
residue. Thus, the methylbilaneenzyme complex really contains a linear
hexapyrrole. The subsequent reaction step, also catalyzed by
porphobilinogen deaminase (Step 5), is the hydrolysis of the bond
linking the second and third pyrrole units of the hexapyrrole to yield
hydroxymethylbilane and the dipyrromethane cofactor. This cofactor
is still linked to the enzyme, which is therefore ready to catalyze a new
round of hydroxymethylbilane synthesis.
 How is the dipyrromethane cofactor assembled?
Porphobilinogen deaminase synthesizes its own cofactor from two PBG
units using, it appears, the same catalytic machinery with which it
synthesizes methylbilane. However, the enzyme Cys reacts much more
rapidly with presynthesized hydroxymethylbilane to form a reaction
intermediate (the product of Step 2) that continues to add two more
PBG units. When hydroxymethylbilane is released, the enzyme retains
its dipyrromethane cofactor.

Cyclization of the hydroxymethylbilane product requires

uroporphyrinogen III synthase. In the absence of this enzyme,
hydroxymethylbilane is released from the synthase and rapidly cyclizes
nonenzymatically to the symmetric uroporphyrinogen I.
Heme biosynthesis takes place partly in the mitochondrion and partly in
the cytosol. ALA is mitochondrially synthesized and is transported to the
cytosol for conversion to PBG and then to uroporphyrinogen III.
Protoporphyrin IX, to which Fe is added to form heme, is produced from
uroporphyrinogen III in reactions catalyzed by (1) uroporphyrinogen
decarboxylase, which decarboxylates all four acetate side chains (A) to
form methyl groups (M); (2) coproporphyrinogen oxidase, which
oxidatively decarboxylates two of the propionate side chains (P) to vinyl
groups (V); and (3) protoporphyrinogen oxidase, which oxidizes the
methylene groups linking the pyrrole rings to methenyl groups. Six
carboxyl groups originally from carboxyl-labeled acetate are lost as
CO2.The only remaining C atoms from carboxyl-labeled acetate are the
carboxyl groups of hemes two propionate side chains (P). During the
coproporphyrinogen oxidase reaction, the macrocycle is transported
back into the mitochondrion for the pathways final reactions.
Protoporphyrin IX is converted to heme by the insertion of Fe(II) into the
tetrapyrrole nucleus by ferrochelatase, a protein that is associated with
the inner mitochondrial membrane on the matrix side.
 The overall pathway of
heme biosynthesis.
Ferrochelatase
catalyzes the insertion
of Fe2+ to yield heme.

-Aminolevulinic acid is synthesized in

the mitochondrion by ALA synthase.
 Mitochondrion

A, P, M, and V, respectively, represent acetyl, propionyl, methyl,

and vinyl (-CH2=CH2) groups. C atoms originating as the carboxyl
group of acetate are red.

ALA leaves the mitochondrion

and is converted to PBG, four
molecules of which condense to
form a porphyrin ring.
The next two reactions involve oxidation of the pyrrole ring substituents yielding
protoporphyrinogen IX whose formation is accompanied by its transport back into the
mitochondrion. After oxidation of the methylene groups linking the pyrroles to yield
protoporphyrin IX.
Heme Biosynthesis Is Regulated Differently in Erythroid and Liver Cells
The two major sites of heme biosynthesis are erythroid cells, which
synthesize 85% of the bodys heme groups, and the liver, which
synthesizes 80% of the remainder. An important function of heme in
liver is as the prosthetic groups of the cytochromes P450, a family of
oxidative enzymes involved in detoxification, whose members are
required throughout a liver cells lifetime in amounts that vary with
conditions. In contrast, erythroid cells, in which heme is, of course, a
hemoglobin component, engage in heme synthesis only on
differentiation, when they synthesize hemoglobin in vast quantities. This
is a onetime synthesis; the heme must last the erythrocytes lifetime
(normally 120 days) since heme and hemoglobin synthesis stop on red
cell maturation (protein synthesis stops on the loss of nuclei and
ribosomes).The different ways in which heme biosynthesis is regulated
in liver and in erythroid cells reflect these different demands: In liver,
heme biosynthesis must really be controlled, whereas in erythroid
cells, the process is more like breaking a dam.
In liver, the main control target in heme biosynthesis
is ALA synthase, the enzyme catalyzing the pathways
first committed step. Heme, or its Fe(III) oxidation
product hemin, controls this enzymes activity
through three mechanisms:
(1) feedback inhibition
(2) inhibition of the transport of ALA synthase (ALAS)
from its site of synthesis in the cytosol to its
reaction site in the mitochondrion
(3) repression of ALAS synthesis.
In erythroid cells, heme exerts quite a different effect on its
biosynthesis. Heme induces, rather than represses, protein
synthesis in reticulocytes (immature erythrocytes). Although
the vast majority of the protein synthesized by reticulocytes is
globin, heme may also induce these cells to synthesize the
enzymes of the heme biosynthesis pathway.
Moreover, the rate-determining step of heme biosynthesis in
erythroid cells may not be the ALA synthase reaction, which
in mammalian reticulocytes is catalyzed by a different isozyme
(ALAS-2) than the ALA synthase that is expressed in other
cells (ALAS-1). Experiments on various systems of
differentiating erythroid cells implicate ferrochelatase and
porphobilinogen deaminase in the control of heme
biosynthesis in these cells. There are also indications that
cellular uptake of iron may be rate limiting.
Iron is transported in the plasma complexed with the iron
transport protein transferrin. The rate at which the iron
transferrin complex enters most cells, including those of liver,
is controlled by receptor-mediated endocytosis. However,
lipid-soluble iron complexes that diffuse directly into
reticulocytes stimulate in vitro heme biosynthesis. The
existence of several control points supports the supposition
that when erythroid heme biosynthesis is switched on, all of
its steps function at their maximal rates rather than any one
step limiting the flow through the pathway. Heme-stimulated
synthesis of globin also ensures that heme and globin are
synthesized in the correct ratio for assembly into hemoglobin.
Seven sets of genetic defects in heme biosynthesis, in liver or erythroid
cells, are recognized. All involve the accumulation of porphyrin and/or
its precursors and are therefore known as porphyrias (Greek: porphyra,
purple). Two such defects are known to affect erythroid cells:
uroporphyrinogen III synthase deficiency (congenital erythropoietic
porphyria) and ferrochelatase deficiency (erythropoietic
protoporphyria). The former results in accumulation of
uroporphyrinogen I and its decarboxylation product
coproporphyrinogen I. Excretion of these compounds colors the urine
red, their deposition in the teeth turns them a fluorescent reddish
brown, and their accumulation in the skin renders it extremely
photosensitive such that it ulcerates and forms disfiguring scars.
Increased hair growth is also observed in afflicted individuals such that
fine hair may cover much of the face and extremities. These symptoms
have prompted speculation that the werewolf legend has a biochemical
basis.
The most common porphyria that primarily affects liver is
porphobilinogen deaminase deficiency (acute intermittent
porphyria).This disease is marked by intermittent attacks of
abdominal pain and neurological dysfunction, often brought
about by infection, fasting, certain drugs, alcohol, steroids,
and other chemicals, all of which induce the expression of
ALAS-1. Excessive amounts of ALA and PBG are excreted
in the urine during and after such attacks. The urine may
become red resulting from the excretion of excess porphyrins
synthesized from PBG in nonhepatic cells although the skin
does not become unusually photosensitive.
 Heme Is Degraded to Bile Pigments
At the end of their lifetime, red cells are removed from the
circulation and their components degraded. Heme catabolism
begins with oxidative cleavage, by heme oxygenase, of the
porphyrin between rings A and B to form biliverdin, a green
linear tetrapyrrole. Biliverdins central methenyl bridge
(between rings C and D) is then reduced to form the red-
orange bilirubin. The changing colors of a healing bruise are a
visible manifestation of heme degradation.
The highly lipophilic bilirubin is insoluble in aqueous
solutions. Like other lipophilic metabolites, such as free fatty
acids, it is transported in the blood in complex with serum
albumin. In the liver, its aqueous solubility is increased by
esterification of its two propionate side groups with
glucuronic acid, yielding bilirubin diglucuronide, which is
secreted into the bile.
Bacterial enzymes in the large intestine hydrolyze the
glucuronic acid groups and, in a multistep process,
convert bilirubin to several products, most notably
urobilinogen. Some urobilinogen is reabsorbed and
transported via the bloodstream to the kidney, where
it is converted to the yellow urobilin and excreted,
thus giving urine its characteristic color. Most of the
urobilinogen, however, is microbially converted to the
deeply red-brown stercobilin, the major pigment of
feces.