L4 Hemoglobin Structure, Synthesis & Breakdown PDF

4 Hemoglobin Structure, synthesis and breakdown ILOs By the end of this lecture, students will be able to 1. 2. 3. 4. 5. Describe normal structure of hemoglobin Describe the pathway for Hb biosynthesis Interpret the regulation of heme synthesis Deduce the end products of Hb catabolism and their rela...

4 Hemoglobin Structure, synthesis and breakdown ILOs By the end of this lecture, students will be able to 1. 2. 3. 4. 5. Describe normal structure of hemoglobin Describe the pathway for Hb biosynthesis Interpret the regulation of heme synthesis Deduce the end products of Hb catabolism and their relation to jaundice Correlate impaired hemoglobin metabolism to clinical outcome ❖ What is Hemoglobin? And why is it important? Hemoglobin (Hb) is a globular protein found exclusively in RBCs. It is responsible for O2 transport from the lungs to the tissues for use in metabolism. Also, it transports protons (H+) and carbon dioxide (CO2) from tissues to the lungs. Hemoglobin is synthesized in progenitors of red blood cells, CANNOT occur in mature RBCs because they lack nucleus. Heme and globin are made separately within the cell before their assembly. Globin is a protein that is transcribed from globin genes, then translated within the ribosomes. Hemoglobin assembly occurs in the cytosol once the globin chains are released from the ribosome. Heme molecules diffuse from the mitochondria into the cytosol, and the globin chains folds around the heme molecules to produce the final hetero-tetramer hemoglobin molecule. Hemoglobin then remains in the cytosol throughout its lifetime.  Hemoglobin structure Hemoglobin molecule (4 heme+ 4 globin) Heme molecule Iron coordination bonds  Hb consists of 4 Heme molecules + 4 Globin chains.  Globin chains are 4 polypeptide subunits, 2 α subunits and 2 β subunits held together by noncovalent interactions.  Heme is a structure consisting of 4 pyrolle rings  Each globin subunit contains a heme moiety with iron atom at its center (one heme/1 globin chain)  Iron in heme moieties is in reduced, ferrous state (Fe2+). Fe2+ binds O2 reversibly  Iron has 6 coordination bonds. Each pyrolle ring in heme is bound to one coordination bond (i.e 4 coordination bonds are consumed by binding to 4 pyrolle rings), the 5 th coordination bond binds to histidine aminoacid in the globin chain and the 6 th coordination bond is left empty to Page 1 of 5 be able to bind to O2 in case of oxygenated hemoglobin, or it can leave the O2 and return unbound in deoxygenated hemoglobin.  This means that each Hb molecule binds 4 O2 molecules, one to each iron molecule present in each of its four heme groups  Hb exists in 2 states Low O2 affinity T (taut/closed) state=Deoxyhemoglobin High O2 affinity R (relaxed/open) state=oxyhemoglobin. R state has 300x greater affinity for O2 N.B. Positive cooperativity of hemoglobin Binding of one O2 molecule to one subunit of deoxyhemoglobin increases affinity for O2 in adjacent subunits, allowing hemoglobin to deliver more O2 to the tissues in response to relatively small changes in the PO2 Sigmoidal shape of oxygen-hemoglobin dissociation curve is characteristic of positive cooperativity Various allosteric effectors influence the equilibrium between the T and R forms and thereby regulate the O2 binding behavior of hemoglobin. Positive effectors that increase oxygen binding to Hb include low CO2, low H+(high alkaline pH), and low concentration of a special metabolite of erythrocytes 2,3-bisphosphoglycerate (BPG). 2,3 BPG is synthesized from 1,3–BPG, an intermediate of glycolysis.(refer to cell biology block, glycolysis) Negative effectors that decrease binding of oxygen and hence facilitate its delivery to tissue are the opposite of positive effectors (see figure) HEME SYNTHESIS Heme is an important biochemical compound that participates in the structure of hemoglobin,cytochrome P450, myoglobin Location of synthesis: involves both the mitochondria and the cytosol. It occurs in nearly every cell, and for Hb synthesis, it occurs in RBC progenitor cells Steps of heme synthesis: Heme is a complex of protoporphyrin IX and ferrous iron (Fe2+). It is synthesized from precursors glycine and succinyl coenzyme A by a series of eight reactions( See figure) Page 2 of 5 Regulation of Heme synthesis ALA synthase is the rate limiting enzyme, it requires vitamin B6 as a coenzyme and is inhibited by heme. When the amount of heme available exceeds the desired amount, heme inhibits ALA synthase in the liver. In this case, Fe2+iron in heme is oxidized to the ferric (Fe3+) state forming hemin. It is hemin that inhibits ALA synthase both by feedback using an allosteric mechanism & also by regulating ALA synthase gene expression by repressing its transcription. Vitamin B6 deficiency also inhibits ALA synthase Lead inhibits both ALA dehydratase & Ferrochelatase Iron deficiency inhibits ferrochelatase Clinical Implications:  Lead poisoning: Induced deficiency in ALA dehydratase and ferrochelatase (↑ in ALA without ↑ in PBG (differentiates it from porphyrias, see later) Presentation: Learning disorders & attention span in children, microcytic anemia,pallor & weakness due to anemia, abdominal pain, lead lines in bone and teeth x-rays, lead lines in gums)  Iron deficiency: Iron is incorporated in the final step of the pathway, Ferrochelatase, which introduces the Fe2+ into the heme ring. Result is microcytic hypochromic anemia.  Vitamin B6 deficiency: Pyridoxine (vit B6) is a cofactor for rate limiting enzyme (ALA synthase). Deficiency is associated with isoniazid therapy for tuberculosis and may cause anemia. Porphyrias: These are inherited defects in heme synthesis, resulting in a heterogeneous group of diseases of porphyrin metabolism. Characterized by a variety of dermatologic, neurologic, and psychological manifestations Symptoms are due to toxic accumulation of pathway intermediates, for example: Aminolevulinic acid (ALA) causes neurological symptoms Protoporphyrins cause photosensitivity. Symptoms are worsened by sunlight and P450 inducing drugs (stimulate the heme synthesis pathway to ↑ production e.g (barbiturates, alcohol) Types Porphyria cutanea tarda  Deficiency in uroporphyrinogen decarboxylase (an adult-onset hepatic porphyria in  Autosomal Dominant  Late onset (4th or 5th decade)  Presentation: Photosensitivity, hyperpigmentation and dark red/brown colored urine Acute intermittent porphyria  Deficiency in porphobilinogen deaminase, ↑ in ALA (δ-aminolevulinic) and↑ in PBG (porphobilinogen)  Autosomal Dominant  Late onset Page 3 of 5  Presentation: NO photosensitivity, episodic psychological symptoms anxiety, depression), abdominal pain and dark red/brown colored urine (paranoia, HEME DEGRADATION (see figure) Steps: (1) Reticulo-endothelial system (RES) RBCs degradation typically occurs around 120 days after maturation, or when cells become abnormal in size or shape. Turnover of RBCs occurs in the reticuloendothelial system (by macrophages of spleen, and liver & BM). Macrophages phagocytose erythrocytes. Hemoglobin is released from degraded RBCs & broken down into heme and globin chains by lysosomal enzymes in macrophages. Globin chains released are broken down into amino acids which are either metabolized or recycled for use in the synthesis of new proteins Heme degradation begins with heme oxygenase, which catalyzes a complex set of reactions that simultaneously open the protoporphyrin ring structure and releases iron in the ferric (Fe3+) state. This results in the conversion of heme to biliverdin (green-colored) Iron (Fe3+) released from Hb in the macrophages is stored bound to ferritin, then transported in the blood by transferrin, which can deliver it to tissues for synthesis of heme. Biliverdin (green color) is then reduced to bilirubin (yellow color) by the cytosolic enzyme biliverdin reductase heme → biliverdin (green-colored) → bilirubin (yellow-colored) (2) Blood Bilirubin is not water-soluble; it is transported in blood bound to serum albumin to liver bilirubin-albumin complex = indirect bilirubin (water insoluble) (3) Liver Bilirubin dissociates from albumin & hepatocytes take up bilirubin Hepatic microsomes conjugate bilirubin with glucoronic acid resulting in conjugated bilirubin = direct bilirubin (water soluble)  A portion of conjugated bilirubin is excreted in urine  Remainder is secreted into bile and then into small intestine (4) Gastrointestinal tract Bilirubin is deconjugated by Intestinal bacteria and metabolized to urobilinogen Majority of urobilinogen is oxidized to stercobilin (orange to yellow) and excreted in feces producing its characteristic color Some of urobilinogen is converted to urobilin and excreted in urine giving its color Some of urobilinogen is absorbed into enterohepatic circulation back to liver Clinical implication o Elevated levels of bilirubin cause jaundice, characterized by yellow skin and sclerae o Accumulation of bilirubin (usually unconjugated) in the basal ganglia of the brain (kernicterus) may result in death or mental retardation if occurs in newborn. Page 4 of 5 Summary of heme degradation Page 5 of 5