MSc Medical Biochemistry,
Hemoglobin is the red blood pigment exclusively
found in erythrocytes.
Concentration of Hb in blood in
Males is 14 – 16 g/dl
Females is 13 – 15 g/dl
Tetrameric allosteric protein
Mol wt 64,450
Conjugated protein globin +heme
Globin is the apoprotein part
Heme is the non-protein part (Prosthetic part)
Example of quaternary structure of protein
Consist of 4 poly peptide chain 2 of one type and 2 of
another type ( α2 and β2) held together by
αhelical conformation constitutes 75% globin
4 of helices are terminated by proline residues.
Each polypeptide chain contains heme group and
oxygen binding cite.
Heme consist of protoporphyrin ring and Fe atom
Fe atom in heme can forms 6 bonds.
4with N of protoporphyrin
one imidazole N of proximal histidine.
In OxyHb , 6th
binds to O2
1.Delivery of O2 from the lungs to the tissues
2. Transport of CO2 and protons (H+
) from tissue to
lungs for expiration.
Effect of 2,3,BPG on O2affinity of Hb
levels are higher in younger children compared to
2,3 BPG is produced in the erythrocytes from an
intermediate (1,3 Bisphosphoglycerate) of glycolysis.
This short pathway is refered as Rapaport Leubering
Significance of Rapaport leubering cycle
It prevents accumulation of ATP not needed by the
It supplies 2,3 BPG in oxygen transport which is
required for the Hb function.
2,3 BPG regulates the binding and release of Oxygen
2,3 BPG accounts for about 16% of the non
carbonate buffer value of erythrocyte fluid.
Iron containing compound belonging to the class of
Protoporphyrin is composed of 4 pyrrole rings which
are linked by methene (=CH) bridges to from
4 methyl , two vinyl and two propionate side chain
gps are attached to the porphyrin rings.
These can be arranged in 15 different ways
One of these isomers called protoporphyrin IX is
) held in the center of the protoporphyrin
molecule by co-ordination bonds with the four
nitrogen of the protoporphyrin ring.
Iron has 6 valency
4 bonds are formed b/n iron and nitrogen atoms
of the porphyrin ring system
bond is formed between nitrogen atom of histidine
residue of the globin polypeptide chain, known as
bond is formed with oxygen
Takes place in all cells (occurs to the greatest extent
in the bone marrow and liver)
Occurs both in mitochondria and cytosol.
Stages of Heme Synthesis
1.Biosynthesis of - aminolevulinic acid (ALA) from
the precursor glycine and succinyl CoA
2.Formation of porphobilinogen (PBG) from δ amino
3. conversion of the porphobilinogen to the cyclic
tetrapyrrol porphyrin ring and heme.
Biosynthesis of δ ALA
step is biosynthesis of Heme
Condensation step (succinyl CoA + glycine)
Rate controlling step in heme synthesis
Occurs in mitochondria
Formation of PBG
Occurs in cytosol
2 mol of δ ALA condense to form one mol of porphobilinogen ad
Enzyme is ALA dehydratase , Zn containing enzyme.
Enzyme is inhibited by lead and ALA is excreted in urine (lead
Conversion of porphobilirogen to porphyrins and
4 BPG condense head to tail to form a linear
Enzyme is PBG deaminase.
PBG cylized spontaneouusly to form UPGI and UPG III ,
Enzyme is UPG III cosynthase
UPG III is converted to CPG III by decarboxylation of all
acetate(A) side chain to methyl (M), enzyme is UPG
CPG III enter mitochondria , it is converted to PPG III by
CPG oxidase enzyme.
This enzyme catalyzed the conversion of two of the
proprionate side chains into vinyl groups by oxidative
This enzyme act only on type III coproporphyrinogen
Oxidation of protoporphyrinogen III to protoporphyrin
is catalyzed by another mitochondrial enzyme,
Final step involves the incoporation ferrous iron into
protoporphyrin in a reaction catalyzed by
mitochondrial heme synthase or ferrochelatase
Regulations of Heme Synthesis
1.ALA synthase allosteric enzyme that catalyzed
controlling step, feed back inhibition by Heme.
2.Iron atom itself may be the active regulatory
3.Several substances like steroid hormone metabolites
ethanol, Barbiturate etc induce the synthesis of
Fate of Bilirlubin
Metabolism & excretion of bilirubin occurs in liver &
Uptake by liver parenchymal cells
Conjugation in smooth endoplasmic reticulum in liver
Secretion of conjugated bilirubin
Excretion of bilirubin in the form of stercobilin and
urobilinogen through feces and urine respectively
Normal concentration of serum bilirubin
Total bilirubin 0.1 - 1.0 mg/dl
Conjugated (direct) bilirubin 0.1 – 0.4 mg/dl
Unconjugated (indirect)bilirubin 0.2 – 0.8 mg/dl
Bilirubin in blood morethan 1mg/dl is called clinical
Hyper bilirubinemia :
Increased bilirubin production
Decreased hepatic uptake
Decreased hepatic conjugation
Decreased excreation of bilirubin into bile
Bilirubin accumulates in allthese conditions
approximately 2.2 - 5 mg/dl.
The skin and sclere appears yellowish due to
deposition of bilirubin in the tissue.
The condition is called jaundice or Icterus.
acquired or inherited
Conjugated or Unconjugated
Hemolytic or prehepatic
Hepato cellular / Hepatic / intrahepatic
Obstructive / Post hepatic
Neonatal / Physiological jaundice
Prehepatic or Hemolytic
eg: - sickle haemoglobin (sickle cell anemia)
- deficiency of enzyme glucose 6-phosphate
- Incompatible blood transfusion
Increased plasma unconjugated bilirubin
Increased amount of urobilinogen in urine and
Absence of bilirubin in urine.
Intrahepatic / Hepatic / Hepatocellular
Infection (viral hepatitis)
Toxic chemicals (alcohol)
Uncojugated hyperbilirubinemia and presence of
some conjugated hyperbilirubinemia
Increased plasma concentrtaion of conjugated and
Decreaed amount of urobilinogen in urine and faeces
Presence of bilirubin in the urine
Raise level of alanine transaminase enzyme
Posthepatic or Obstructive Jaundice
occurs due to obstruction in the passage of
conjugated bilirubin from the liver cells to the intestine
this condition is also called as cholestasis.
Blockage of common bile duct by gallstones
Carcinoma of the head of the pancreas
carinoma of the duct
Increased conjugated bilirubin in plasma
Absence of urobilinogen in faeces and urine
Presence of bilirubin and bile salts in urine
Raised ALP in plasma
Neonatal of physiologic jaundice
Mild jaundice in the first few days after birth
Results from an accelerated haemolysis and
immature liver enzyme system for conjugation
Liver is deficient in enzyme UDP-glucuronyl
Increased unconjugated bilirubin is seen
If it exceeds 20 – 25 mg/dl it penetrates the BBB
This results in hyperbilirubinaemic toxic
encephalopathy or kernicterus