1. David Hart Dec 12, 2006

2. Heme

3. Porphyrins Cyclic compounds that bind metal ions Chlorphyll (Mg 2+ ) –Central to solar energy utilization Heme (Fe 2+ ) –Most prevalent metalloporphyrin in humans –Central to oxygen sensing and utilization Cobalamin (Cobalt)

5. The Heme Pocket in Hemoglobin

6. Heme One ferrous (Fe 2+ ) atom in the center of the tetrapyrrole ring of Protoporphyrin IX Prosthetic group for –Hemoglobin and Myoglobin –The Cytochromes –Catalase and Tryptophan pyrrolase –Nitric Oxide Synthase Turnover of Hemeproteins (Hemoglobin, etc) is coordinated with synthesis and degradation of porphyrins Bound iron is recycled

7. Lecture Outline Heme function Heme synthesis and regulation Iron metabolism Porphyrias Heme degradation

8. Heme Function Oxygen sensing (heme and hemoproteins) Oxygen transport (hemoglobin) Oxygen storage (myoglobin) Electron transport (cytochromes) Oxidation (cyrochrome p450, tryptophan pyrrolase, guanylate cyclase …) Decomposition and activation of H 2 O 2 (catalase and peroxidase) Nitric Oxide Synthesis Regulation of cellular processes Effector of apoptosis

9. N NHN HN B C D Porphyrin: Cyclic molecule formed by linkage of four pyrrole rings through methenyl bridges A

10. Porphyrin Side Chains M = Methyl (-CH 3 ) V = Vinyl (-CH=CH 2 ) P = Propionyl (-CH 2 -CH 2 -COO - ) A = Acetyl (-CH 2 -COO - )

11. Biosynthesis of Heme Synthesized in every human cell Liver (15%): –65% Cytochrome P450 –Synthesis fluctuates greatly –Alterations in cellular heme pool Bone Marrow (80%) –Erythrocyte precursors: Hemoglobin –Synthesis relatively constant –Matched to rate of globin synthesis –Largely unaffected by other factors

12. COOH CH 2 COSCoA CH 2 NH 2 COOH SUCCINYL CoA GLYCINE All Carbon and Nitrogen atoms provided by 2 building blocks:

13. COOH CH 2 COSCoA CH 2 NH 2 COOH SUCCINYL CoA GLYCINE is Decarboxylated IN MITOCHONDRIA  AMINOLEVULINIC ACID SYNTHASE - CO 2

14. COOH CH 2 C=O CH 2 NH 2 Condense to form:  AMINOLEVULINIC ACID (  ALA) MOVES OUT OF THE MITOCHONDRION

15. COOH CH 2 C=O CH 2 NH 2 COOH CH 2 C=O CH 2 NH 2 2 Molecules dehydrated by  ALA DEHYDRATASE -2 H 2 O

16. COOH CH 2 C NH COOH CH 2 C CH 2 NH 2 To form Porphobilinogen (PBG)

17. COOH CH 2 NHNH COOH CH 2 NH 2 Porphobilinogen (PBG) Acetate CH 2 COO - Propionate CH 2 CH 2 COO -

18. Porphobilinogen (PBG) NHNH CH 2 NH 2 A P

19. NHNH CH 2 NH 2 A P NHNH CH 2 NH 2 A P NHNH CH 2 NH 2 A P NHNH CH 2 NH 2 A P

20. Hydroxymethylbilane synthase & Uroporphyrinogen III synthase Four PBG molecules condense Ring closure Isomerization

21. NH HN A B C D A A P A P P P A Uroporphyrinogen III

22. NH HN HOOC-H 2 C- -CH 2 -CH 2 -COOH -CH 2 -COOH CH 2 COOH CH 2 COOH CH 2 COOH CH 2 Uroporphyrinogen III

23. Series of decarboxylations & oxidations Porphyrinogens: –Chemically reduced –Colorless intermediates Porphyrins: –Intensely colored –Fluorescent Uroporphyrinogen III  Coproporphyrinogen III  Moves back into Mitochondrion Protoporphyrinogen IX  Protoporphyrin IX

24. NH NHN N H 3 C- -CH=CH 2 -CH 3 CH 2 COOH CH 2 COOH CH 3 Protoporphyrin IX CH=CH 2

25. HEME Fe 2+ chelated by Protoporphyrin IX Assisted by Ferrochelatase CH 3 -

26. Regulation of Heme Synthesis

27.  AMINOLEVULINIC ACID SYNTHASE Two tissue-specific isozymes Coded on separate genes In Liver, heme represses synthesis and activity of  ALAS –Heme can be used for treatment of acute porphyric attack In RBC heme synthesis regulation is more complex –Coordinated with globin synthesis

28. COOH CH 2 COSCoA CH 2 NH 2 COOH SUCCINYL CoA GLYCINE IN MITOCHONDRIA  AMINOLEVULINIC ACID SYNTHASE RATE-CONTROLLING STEP IN HEPATIC HEME SYNTHESIS COOH CH 2 C=O CH 2 NH 2  ALA

29. Bonkovsky ASH Education Book December 2005

30. Disorders of Heme Synthesis X-linked Sideroblastic Anemia Lead Poisoning Iron Deficiency Anemia The Porphyrias

31. X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia  ALAS Requires Pyridoxal Phosphate as Coenzyme Some Sideroblastic Anemias improve with Pyridoxine (B 6 )

32. COOH CH 2 C=O CH 2 NH 2 COOH CH 2 C=O CH 2 NH 2 -2 H 2 O  ALA moves out of the mitochondrion  ALA DEHYDRATASE Inhibited by Heavy Metal: LEAD POISONING PBG NHNH CH 2 NH 2 A P

33. Lead Poisoning

34.  ALAD and Ferrochelatase Are particularly sensitive to Lead inhibition Lead Poisoning Fe + PPIX Ferrochelatase Heme

35. Iron Metabolism Reactive Transition Metal (Fe 2+  Fe 3+ ) Normally present complexed with proteins that limit its reactivity Both iron deficiency and iron overload cause cellular defects and disease Most available iron generated by macrophages that recycle red cell iron Dietary Fe 3+ in duodenum converted to Fe 2+ and absorbed by duodenal enterocyte

36. Iron 35% of Earth’s mass nasa

37. Fe 3+ Heme Fe 2+ Blood Apical Duodenal Enterocyte Mitochondrial Heme Synthesis Hepatocyte Macrophage Erythroid Cell diFe 3+ Transferrin GUT Contents

38. NEJM June 2004

39. Fe 2+ Blood Macrophage RBC Hemoglobin Haptoglobin Heme Hemopexin

40. Syed, Hemoglobin 2006 ?

41. http://walz.med.harvard.edu

42. Hentze, Muckenthaler & Andrews Cell, Vol 117, 285-297, April 30, 2004 Hepcidin

43. Hepcidin: 25 Amino Acids J Med Genet 2004

44. Nemeth et al, Science, Dec 2004

45. Beutler, Science Dec 2004

46. Hentze, Muckenthaler & Andrews Cell, Vol 117, 285-297, April 30, 2004 Ferroportin

47. Genetic Hemochromatosis Disruption of Hepcidin / Ferroportin Autosomal Recessive –HFE C282Y/C282Y –TfR2 –Hemojuvelin –Hepcidin Autosomal Dominant –Ferroportin

48. medlib.med.utah.edu Normal Liver

49. www.med.niigata-u.ac.j Granular, Dark Reddish Brown Surface of Liver in Hemochromatosis

50. http://eduserv.hscer.washington.edu Iron Accumulation in Chronic Disease

51. Ring Sideroblast Prussian Blue stains Iron In Mitochondria www.uchsc.edu

52. Iron Deficiency Anemia Hypochromic, Microcytic

53. http://eduserv.hscer.washington.edu Normal Red Blood Cells

55. www.lsuagcenter.co Spinach: Non-Heme Iron Less Readily Absorbed Oxalates Phytates Tannins Fiber Calcium

56. www.mcgil.com/food/pics Heme Iron is More Readily Absorbed

57. www.agnet.org/library Iron Deficient Spinach “Chlorosis”

58. www.geoimagery.com Harvesting Latex

59. www.sentientkinetics.com Geophagia

60. www.awesomedrinks.com Pagophagia

61. Solemnity Scale: 0 = No smiles/hour 5 = “wreathed” In smiles

62. www.drmhijazy.com Spoon Nails

63. Blue Sclera

64. Disorders of Heme Synthesis X-linked Sideroblastic Anemia Lead Poisoning Iron Deficiency Anemia The Porphyrias

65. Heme  porphuros (purple)

66. Heme Synthesis: Porphyrias 8 Enzymatic Reactions 7 Deficiencies: “Porphyrias” Most are Autosomal Dominant Hepatic or Erythroid depending on main site of synthesis / accumulation

67. Porphyrias Accumulation and excretion of porphyrins –Pattern depends on which enzyme affected Multiple alleles Acute and Chronic –Acute: Neurovisceral attacks Porphyrin accumulation: Photosensitivity –Formation of reactive oxygen species –Damage tissues, Release lysosomal enzymes

68. Very Rare Recessive Porphyria ALA-D Porphyria Lead Poisoning

69. Acute Hepatic PBG and  ALA Accumulate in Urine PBG in Urine: Diagnostic Screen Urine darkens with exposure NOT photosensitive Neuro-visceral attacks Precipitated by Drugs, EtOH which induce cytochrome P450 Hydroxymethylbilane Synthase Lead Poisoning ALA-D Porphyria