HEAMOGLOBIN

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Hemoglobin

• M.Prasad Naidu
• MSc Medical Biochemistry,
• Ph.D.Research Scholar

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Hemoglobin (Hb)

• Hemoglobin is the red blood pigment exclusively
  found in erythrocytes.
• Normal range
• Concentration of Hb in blood in
• Males is 14 16 g/dl
• Females is 13 15 g/dl

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• Structure
• Tetrameric allosteric protein
• Mol wt 64,450
• Conjugated protein globin heme
• Globin is the apoprotein part
• Heme is the non-protein part (Prosthetic part)
• Chromo protein
• Example of quaternary structure of protein
• Globular protein
• Consist of 4 poly peptide chain 2 of one type and
  2 of another type ( a2 and ß2) held together by
  noncovalent bonds.

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• ahelical conformation constitutes 75 globin
  molecules.
• 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
• 5th one imidazole N of proximal histidine.
• In OxyHb , 6th binds to O2

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• Function
• 1.Delivery of O2 from the lungs to the tissues
• 2. Transport of CO2 and protons (H) from tissue
  to
• lungs for expiration.

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• Effect of 2,3,BPG on O2affinity of Hb
• levels are higher in younger children compared
  to elders.
• 2,3 BPG is produced in the erythrocytes from an
  intermediate (1,3 Bisphosphoglycerate) of
  glycolysis.
• This short pathway is refered as Rapaport
  Leubering cycle.

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• Significance of Rapaport leubering cycle
• It prevents accumulation of ATP not needed by
  the erythrocyte
• 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 from hemoglobin.
• 2,3 BPG accounts for about 16 of the non
  carbonate buffer value of erythrocyte fluid.

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• HEME
• Iron containing compound belonging to the class
  of protoporphyrin compounds
• Protoporphyrin is composed of 4 pyrrole rings
  which are linked by methene (CH) bridges to from
  tetrapyrrole (porphyrin)
• 4 methyl , two vinyl and two propionate side
  chain gps are attached to the porphyrin rings.

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• These can be arranged in 15 different ways
• One of these isomers called protoporphyrin IX is
  biologically active.
• Iron (Fe2) held in the center of the
  protoporphyrin molecule by co-ordination bonds
  with the four nitrogen of the protoporphyrin
  ring.

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• Iron has 6 valency
• 4 bonds are formed b/n iron and nitrogen atoms
  of the porphyrin ring system
• 5th bond is formed between nitrogen atom of
  histidine residue of the globin polypeptide
  chain, known as proximal histidine.
• 6th bond is formed with oxygen

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• Heme Synthesis
• Takes place in all cells (occurs to the greatest
  extent in the bone marrow and liver)
• Occurs both in mitochondria and cytosol.

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• Stages of Heme Synthesis
• 1.Biosynthesis of - aminolevulinic acid (ALA)
  from the precursor glycine and succinyl CoA
• 2.Formation of porphobilinogen (PBG) from d amino
  levulinic and
• 3. conversion of the porphobilinogen to the
  cyclic tetrapyrrol porphyrin ring and heme.

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• Biosynthesis of d ALA
• 1st step is biosynthesis of Heme
• Condensation step (succinyl CoA glycine)
• Rate controlling step in heme synthesis
• Occurs in mitochondria

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• Formation of PBG
• Occurs in cytosol
• 2 mol of d ALA condense to form one mol of
  porphobilinogen ad 2H2O.
• Enzyme is ALA dehydratase , Zn containing enzyme.
• Enzyme is inhibited by lead and ALA is excreted
  in urine (lead poisoning).

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• Conversion of porphobilirogen to porphyrins and
  heme
• 4 BPG condense head to tail to form a linear
  tetrapyrrole.
• 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 decarboxylase.
• CPG III enter mitochondria , it is converted to
  PPG III by CPG oxidase enzyme.

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• This enzyme catalyzed the conversion of two of
  the proprionate side chains into vinyl groups by
  oxidative decarboxylation.
• This enzyme act only on type III
  coproporphyrinogen
• Oxidation of protoporphyrinogen III to
  protoporphyrin is catalyzed by another
  mitochondrial enzyme, protoporphyrinogen oxidase.

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• Final step involves the incoporation ferrous
  iron into protoporphyrin in a reaction catalyzed
  by mitochondrial heme synthase or ferrochelatase

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• 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
  species
• 3.Several substances like steroid hormone
  metabolites ethanol, Barbiturate etc induce the
  synthesis of hepatic ALA

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• Disorders of Heme biosynthesis
• Porphyrias

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• Breakdown of Hemoglobin
• RBC life span is 120 days , after this they are
  degraded by RE system

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Fate of Bilirubin
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• Fate of Bilirlubin
• Metabolism excretion of bilirubin occurs in
  liver intestine
• 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

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Disorders of Hemoglobin Catabolism

• 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 hyperbilirubinemia.

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• 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.

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• Jaundice (Hyperbilirubimia)
• acquired or inherited
• Conjugated or Unconjugated
• 4 types
• Hemolytic or prehepatic
• Hepato cellular / Hepatic / intrahepatic
• Obstructive / Post hepatic
• Neonatal / Physiological jaundice

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• Acquired hyperbilirubimia
• Prehepatic or Hemolytic
• eg - sickle haemoglobin (sickle cell anemia)
• - deficiency of enzyme glucose 6-phosphate
  dehydrogenase.
• - Incompatible blood transfusion
• Biochemical features
• Increased plasma unconjugated bilirubin
• Increased amount of urobilinogen in urine and
  Feces.
• Absence of bilirubin in urine.

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• Intrahepatic / Hepatic / Hepatocellular Jaundice
• Infection (viral hepatitis)
• Toxic chemicals (alcohol)
• Drugs
• cirrhosis
• Uncojugated hyperbilirubinemia and presence of
  some conjugated hyperbilirubinemia
• Biochemical Features
• Increased plasma concentrtaion of conjugated and
  unconjugated bilirubin.
• Decreaed amount of urobilinogen in urine and
  faeces
• Presence of bilirubin in the urine
• Raise level of alanine transaminase enzyme

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• 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.
• Causes
• Blockage of common bile duct by gallstones
• Carcinoma of the head of the pancreas
• carinoma of the duct

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• Biochemical Features
• Increased conjugated bilirubin in plasma
• Absence of urobilinogen in faeces and urine
• Presence of bilirubin and bile salts in urine
• Raised ALP in plasma

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• 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
  transferase.
• Increased unconjugated bilirubin is seen
• If it exceeds 20 25 mg/dl it penetrates the BBB
• This results in hyperbilirubinaemic toxic
  encephalopathy or kernicterus

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• Inherited hyperbilirubinaemia
• Gilberts syndrome
• Crigler Najjar syndrome
• Dubin Johnson syndrome
• Rotor syndrome

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