Hemoglobin and Myoglobin

Cont-

heme containing
Myoglobin, monomeric protein of red muscle
(heart and skeletal) , stores oxygen as a
reserve against oxygen deprivation. 153 aa,
17,200 MW;
Exhibits Michaelis-Menten properties

Hemoglobin, a tetrameric protein of
erythrocytes, transports O2 to the tissues and
returns CO2 and protons to the lungs. two (α )
alphas of 141 residues, 2 (β)betas of 146
Exhibits allosteric properties



Hemeproteins
 are a group of specialized proteins that
contain heme as a tightly bound prosthetic
group. Oxygen Can Be Bound to a Heme
Prosthetic Group
They maintain a supply of oxygen essential for

oxidative metabolism
Proteins
Composition
Classification
 Structural
 Functional



 HEME-CONTAINING PROTEINS

 Cytochromes = involves in electron

transport chain.
 Catalase = degradation of hydrogen
peroxide
•Tryptophan pyrolase = oxidation of
tryptpphan
•Ctochrome P450 = hydroxylation of
xinobiotics
 Hemoglobin =
•Myoglobin =
Heme synthesis
PORPHYRINS and IRON
are cyclic compounds
formed by the linkage of
four pyrrole rings
through –HC = methenyl
bridges
 porphyrias = caused by
abnormalities in the pathway of
biosynthesis, not very
prevalent
 Jaundice bilirubin in the, due to
overproduction or to failure of
its excretion.
 A characteristic property of the
porphyrins is the formation of
complexes with metal ions
bound to the nitrogen atom of
the pyrrole rings
Oxygen transport
Haemoglobin is a
conjugated protein
attached to a
prosthetic group
The prosthetic group
is called haem and
contains iron
There are four haem
groups in each
haemoglobin
molecule each of
which can bind to
an oxygen
molecule.
Myoglobin
 Single polypeptide
 a monomeric heme protein
 16,700 daltons Rich in α
Helix
 8 α helices (A-H)
 Located in skeletal & Heme prosthetic group
cardiac muscle. high in
diving mammals like
whale & seals
 Fe in Mb is Fe2+ - ferrous
iron - the form that binds
oxygen
 Oxidation of Fe yields 3+
charge - ferric iron
-metmyoglobin does not
bind oxygen http://www.agen.ufl.edu/~chyn/age2062/lect/lect_02/3_27.gif

 Oxygen binds as the sixth

ligand to Fe
The Conformation Change

 The secret of Mb and Hb!

Oxygen binding changes the Mb
conformation
Without oxygen bound, Fe is out of heme
plane
Oxygen binding pulls the Fe into the heme
plane
Fe pulls its His F8 ligand along with it
The F helix moves when oxygen binds
Total movement of Fe is 0.029 nm - 0.29 A
This change means little to Mb, but lots to
 Heme Prosthetic Group
Heme (Fe2+ ) has
affinity for O2.

N
Hematin (Fe3+ )
N Fe N cannot bind O2.

N
Located in crevice
HO
O where it is
protected from
oxidation.
 Oxygen Binding to
Myoglobin
d ista lh istid in e
O2 binds to only
available
coordination site on
iron atom.

His 93 (proximal his)
binds directly to iron.

His 64 (distal his)
stabilizes the O2
binding site.
p roxim a l h istid in e

http://cwx.prenhall.com/horton/medialib/media_portfolio/text_images/FG04_44.JPG
 O2 Binding Curve
Myoglobin has high 100
affinity for O2.


a rte ria lp re ssu re

saturation with
P50 = 2.8 Torr

tissu e s
 50
Allows myoglobin to
act as O2 storage
reserve. O2

Releases O2 when pO2 2.8 20 100
becomes low pO2 (partial pressure of
indicating O2
O2) (Torr)
deprivation.
HEME & FERROUS IRON CONFER THE ABILITY
TO STORE & TO TRANSPORT OXYGEN
heme, a cyclic tetrapyrroleconsisting of four
molecules of pyrrole linked by α-methylene
bridges.
This planar network of conjugated double
bonds absorbs visible light and colors heme
deep red.

Heme synthesis
PORPHYRINS and IRON
are cyclic compounds
formed by the linkage of
four pyrrole rings
through –HC = methenyl
bridges
 porphyrias = caused by
abnormalities in the pathway of
biosynthesis, not very
prevalent
 Jaundice bilirubin in the, due to
overproduction or to failure of
its excretion.
 A characteristic property of the
porphyrins is the formation of
complexes with metal ions
bound to the nitrogen atom of
the pyrrole rings
S tru ctu r
e

N
Pyrrole
Iron Utilization
 N a tu ra l P o rp h y rin s h a v in g S u b stitu e n t S id e C h a in s
 Fischer shorthand formula
 methenyl bridges are omitted
and each pyrrole ring is
shown with the eight
substituent positions
 The arrangement of the acetate
(A) and propionate (P)
substituents in the
uroporphyrin (in ring IV, the U ro p o rp h yrin III. A ( a ce ta te ) =
expected order of the A and P -CH2COOH ; P ( propionate ) =
substituents is reversed). -CH2CH2COOH
 this type of asymmetric
substitution is classified
as a type III porphyrin.
 A completely symmetric
arrangement = as a type I
porphyrin.
 Only types I and III are found in
nature
 type III series is more abundant
and important b/c includes
heme .
Heme
synthesis
STARTING MATERIAL
B6
RATE LIMITING
ENZYME
 O
− −
OOC CH2 CH2 C S-CoA + OOC CH2 NH3+
succinyl-CoA H+ glycine
δ-Aminolevulinic
Acid Synthase CoA-SH CO2
O
−
OOC CH2 CH2 C CH2 NH3+
δ-aminolevulinate (ALA)

Heme synthesis begins with



condensation of glycine & succinyl-CoA,

with decarboxylation, to form δ -
aminolevulinic acid (ALA).
 H O
Pyridoxal phosphate O− C
−O
(PLP) serves as H2
P C OH
coenzyme for δ - O
Aminolevulinate O
Synthase (ALA +
Synthase), an enzyme N CH3
H
evolutionarily related to Pyridoxal phosphate (PLP)
transaminases.
Condensation with

succinyl-CoA takes H2C COO−

place while the amino N+
group of glycine is in O− HC H
−O
Schiff baselinkage to P
H2
C O−
the PLP aldehyde. O
CoA & the glycine O
carboxyl are lost +
N CH3
following the H
condensation. glycine-PLP Schiff base (aldimine)
ALA Synthase is the committed step of the
heme synthesis pathway, & is usually rate-
limiting for the overall pathway.
Regulation occurs through control of gene

transcription. 
Heme functions as a feedback inhibitor,

repressing transcription of the ALA

Synthase gene in most cells.
A variant of ALA Synthase expressed only in

developing erythrocytes is regulated instead by

availability of iron in the form of iron-sulfur
clusters.
 Cytosol
two molecules of ALA are condenses
ALA dehydratase is a zinc-containing
enzyme
is sensitive to inhibition by lead ( lead
poisoning.)
The Zn++ binding sites in the homo-octomeric
mammalian Porphobilinogen Synthase, which
include cysteineS ligands, can also bind Pb++
(lead).
Inhibition of Porphobilinogen Synthase by Pb++

results in elevated blood ALA, as impaired

heme synthesis leads to de-repression of
transcription of the ALA Synthase gene. High
ALA is thought to cause some of the neurological
−
++ COO−
effects of lead poisoning, although Pb COO
also
may directly affect the nervous system.CH2 CH2
ALA is toxic to the brain, perhaps due CH2 to: CH2
• Similar ALA & neurotransmitter GABAC O CH2
(γ -aminobutyric acid) structures.CH2 NH3+
• ALA autoxidation generates reactiveNH3+
oxygen species (oxygen radicals).ALA GABA
 COO−

COO− CH2

CH2 CH2
P o rp h o b ilin o g e
n ( P B G ) is the
H2C
N
first p a th w a y N
H in te rm e d ia te th a t NH3+ H
pyrrole in clu d e s a Porphobilinogen (PBG)
p y rro le ring.
T h e p o rp h y rin ring is formed by
co n d e n sa tio n o f 4 molecules of
p o rp h o b ilin o g e n . 
P o rp h o b ilin o g e n D e a m in a se ca ta lyze s
su cce ssive P B G co n d e n sa tio n s, initiatedin
e a ch ca se b y e lim in a tio n o f th e amino
Disorders of Heme Synthesis

Acquired: Lead poisoning


Congenital: Porphyrias

Deficiency of heme has far-reaching effects
(hemoglobin, cytochromes, etc.)



LEAD TOXICITY
Symptoms
 Irritibility  Poor appetite
 Lethargy  Abdominal pain (with or
 Sleeplessness without vomiting)
 Headaches  Constipation

Pathophysoiology
 Binds to any compound with a sulfhydryl group
 Inhibits multiple enzyme reactions including
those involved in heme biosynthesis (ALA synthase &
ferrochelatase)
 One symptom of lead toxicity is increases in 5-
ALA without concomitant increases in PBG
HEME SYNTHESIS (CONT.)

Vitamin
B6

lead
 PORPHYRIAS
 A group of rare disorders caused by deficiencies of enzymes of
the heme biosynthetic pathway

•The majority of the porphyrias are inherited in a autosomal

dominant fashion - thus, affected individuals have 50% normal
levels of the enzymes, and can still synthesize some heme
•
•Affected individuals have an accumulation of heme precursors
(porphyrins ), which are toxic at high concentrations
•
•Attacks of the disease are triggered by certain drugs, chemicals,
and foods, and also by exposure to sun

 Treatment involves administration of hemin, which provides

negative feedback for the heme biosynthetic pathway, and therefore,
prevents accumulation of heme precursors
Scriver et al., The Metabolic & Molecular Basis of Inherited Disease, 8th edition, 2001.
ACUTE INTERMITTENT PORPHYRIA

•Hepatic, autosomal dominant

•Caused by a deficiency in porphobilinogen

deaminase, which is involved in the conversion of
porphobilinogen (PBG) to uroporphyrinogen III

•PBG, uroprophryin, and 5-ALA accumulate in the plasma

and the urine

•Patients have neuropyschiatric symptoms and

abdominal pain (neurovisceral)
PORPHYRIA CUTANEA TARDA
•Most common porphyria
•
•Hepatic, autosomal dominant

•Disease is caused by a deficiency in uroporphyrinogen

decarboxylase, which is involved in the conversion of
uroporphyrinogen III to coproporphyrinogen III

 Uroporphyrinogen accumulates in urine

 Patients are photosensitive (cutaneous photosensitivity)

Accumulation of porphyrinogens results in their
conversion to porphyrins by light

Porphyrins react with molecular oxygen to form

oxygen radicals

Oxygen radicals can cause severe damage to the

skin