1. Pathophysiology of Heme Synthesis Beth A. Bouchard BIOC 212: Biochemistry of Human Disease Spring 2006

2. HEME-CONTAINING PROTEINS  Hemoglobin  Myoglobin  Cytochromes  Catalase  Some peroxidases

3. STRUCTURE OF HEME

4. An Overview of Iron Metabolism Gut Blood Cells Low pH of stomach solubilizes Fe-containing ionic compounds. Fe transporters facilitate absorption into blood stream Fe 3+ ions are bound and chelated by Transferrin (Tf). Transferrin transports Fe to tissues Maintains solubility Keeps Fe ions unreactive Transferrin endocytosis is receptor-mediated (TfR) Endocytosis results in Fe 3+ release Fe is distributed to topologically distinct regions of the cell via Fe transporter and/or channels (?) Usage: Protein components (Heme) Storage: Ferritin (Fe 2+ ) Toxicity

5. Roles of Iron in the Cell Transferrin Receptors (TfR) Fe(III) 2 -Tf Tf Proteins: Catalysis Electron, oxygen transport Structural stabilization Sensor of Fe, ROS Formation of protein-bound radicals Ferritin Storage and Sequestration: Ferritin Toxicity: Oxidative stress [Fe]

6. Iron Control of Translation IREs are found in the 5’-UTR or the 3’- UTR of mRNAs Regulate mRNA translation via IRBP Decreased cellular iron levels: –IRBP is free of iron and can therefore, interact with the IREs in the 3'-UTR of the transferrin receptor (TfR) mRNA, which prevents its degradation. –IRBP binds to the IRE in the 5’-UTR of the ferritin mRNA preventing its translation. Increased cellular iron levels: –IRBP binds iron and cannot interact with the IREs in the TfR mRNA resulting in an increase in its degradation. –IRBP cannot bind to the IRE in the ferritin mRNA allowing for its translation. IRE

7. STRUCTURE OF HEME Ferrous iron (Fe 2+ ) Protoporphyrin IX: contains 4 pyrrole rings linked together by methenyl bridges

8. Heme 8 8 Succinyl CoA Glycine** HEME SYNTHESIS ** Amino acid (building blocks of protein) synthesized in your body

9. Heme synthesis

10. HEME SYNTHESIS

11. HEME SYNTHESIS: Red blood cells 85% of total heme synthesis occurs in red blood cells (RBC) Ceases when RBC’s mature Erythroid-specific ALA synthase is regulated by an IRE in the mRNA – binding of IRBP inhibits mRNA translation

12. Heme stimulates hemoglobin synthesis in reticulocytes

13. HCI = heme controlled inhibitor Reduced initiation of translation

14. ** * In RBCS, heme synthesis is also regulated at the level of the en- zymes ferrochelatase* and porphobilinogen deaminase**

15. HEME SYNTHESIS: Liver The liver is the main non-RBC source of heme synthesis Heme produced in the liver is used mainly for the synthesis of the cytochrome P 450 class of enzymes that are involved in detoxification Regulated at level of ALA synthase: Formation of 5- ALA is the rate-limiting step in heme synthesis in the liver

16. Formation of 5-aminolevulinate (5-ALA) 5-ALA  5-ALA is formed in the mitochondria and transported to the cytoplasm

17. Regulation of ALA Synthase Level of enzyme synthesis Enzyme synthesis, as well as its transport to the mitochondria, is inhibited by elevated levels of heme and hemin, the Fe 3+ oxidation product of heme Enzyme synthesis is upregulated by a large number of drugs including barbiturates, steroids with a 4,5 double bond (e.g. testosterone) and some oral contraceptives: These drugs are metabolized by the microsomal cytochrome P 450 mono-oxygenase system, a heme- containing protein. Level of enzyme activity Heme and hemin inhibit ALA synthase activity Requires pyridoxal phosphate (Vitamin B6) as a coenzyme

18. Disorders of Heme Synthesis Acquired: Lead poisoning Congenital: Porphyrias Deficiency of heme has far-reaching effects (hemoglobin, cytochromes, etc.)

19. LEAD TOXICITY Symptoms  Irritibility  Poor appetite  Lethargy  Abdominal pain (with or  Sleeplessnesswithout vomiting)  Headaches  Constipation Pathophysoiology  Binds to any compound with a sulfhydryl group  Inhibits multiple enzyme reactions including those involved in heme biosynthesis (PBG synthase & ferrochelatase)  One symptom of lead toxicity is increases in 5-ALA without concomitant increases in PBG

20. HEME SYNTHESIS (CONT.) lead Vitamin B6

21. 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

23. Scriver et al., The Metabolic & Molecular Basis of Inherited Disease, 8 th edition, 2001.

24. 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)

25. 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