2011/2012 2. semester Biochemistry (Cell and Organ Biochemistry) 2.

lecture Biochemistry of Iron y Metabolism

nd nd

2012/02/14

Dr Rka Tth Rvszn Biochemistry and Molecular Biology Department y gy p Lectures for 2nd year Physiotherapyst
(based partly on slides of Pter Bagossi and slides from )

BIOCHEMISTRY
COMPULSORY READING: Lecture presentations with short explanations are available on the web page of the department: (http://bmbi.med.unideb.hu). p ( p ) Username: student , password: student2011.
Downloads/ Educational materials in English/ Physiotherapists/ Biochemistry FURTHER READINGS: Biochemistry and Molecular Biology Syllabus III. (ed .by Prof Lszl Fss) chapter 5.1. Devlin: Textbook of Biochemistry (5thed Wiley) chapter 24 Iron and heme metabolism (105324. 1075.p) Harvey, Ferrier: Biochemistry 6th ed. (Lippincott, 2011) chapter 21. Haem metabolism

Supplementary Most important obligatory

CONTENTS
I.
1. 2. 2 3. 4.

IRON METABOLISM
Introduction Absorption of iron Transport of iron and Storage of iron g p Regulation of iron metabolism: hepcidin

II.
1. 2.

HEME METABOLISM
Biosynthesis of heme, Porphyrias Degradation of heme, Jaundice

III. HEMOGLOBIN, MYOGLOBIN

1. 2. 3. 3 4. Structure of hemoglobin Polymorphism of globins O2 binding and its regulation by 2 3BPG pH, CO2, temperature 2,3BPG, pH CO2 Abnormal hemoglobins: Sicle cell anemia, MetHb, HbA1c

IRON METABOLISM

Major problems of iron metabolism

Iron is an essential metal for humans, involved in metabolism and transport of oxygene but free iron is dagerous both iron deficiency anaemia (affects over 30% of the worlds population) and h ld l ti ) d hemochromatosis (i h t i (iron overload) l d) are dangerous Humans regulate iron homeostasis only through control of iron absorption from the diet
5

Iron is involved in the metabolism and transport of Oxygen

Iron containing proteins: I t i i t i Hem containing proteins
Oxygen transport and storage -hemoglobin -myoglobin Electrons transporters- cytochromes in the electron transport chain O2 activators Cytochrome P450 enzymes NADPH oxydase Tryptophan pyrrolase Catalases d C t l degrade H2O2 d NO synthetase FeS cluster proteins (electron transport,succinate DH, aconitase) Iron in the catalytic centre various oxidoreductases (RR, Homogentisate oxidase, Lys, Pro, Phe, Tyr hydroxilase) Proteins in the transport and storage of iron (ferritin transferrin (ferritin,transferrin, lactoferrin)
6

Iron is dangerous
free iron
generates reactive oxygen species (H2O2 2OH-) causes distorsion in the structure of macromolecules forms complexes with anions, which are precipitated used by bacteria

Solution: iron is tightly bound to proteins in the body

7

Human iron metabolism Iron metabolism is the set of chemical reactions maintaining human homeostasis of iron. Iron is an absolute requirement for most forms of life, including humans and most bacterial species. Because plants and animals all use iron, iron can be found in a wide variety of food sources (meat, liver, dried leguminoses, dried fruits, fortified flour, cereals). Iron is essential to life because of its unique ability to serve as both an electron donor and life, acceptor. However, iron can also be potentially toxic. Its ability to donate and accept electrons means that if iron is free within the cell, it can catalyze the conversion of hydrogen peroxide into free radicals (Fenton reaction). Free radicals can cause damage to a wide variety of cellular structures, and ultimately kill the cell. In addition, free iron causes distorsion in the structure of macromolecules and forms complexes with anions, which are precipitated within the cells. To prevent that kind of damage, all life forms that use iron, bind the iron ions to proteins That allows the cells to use the benefits of iron but also limit its proteins. iron, ability to do harm. Iron containing proteins Most well-nourished people in industrialized countries have 3-4 grams of iron in their bodies. Of this, about 2.5 g is contained in the hemoglobin needed to carry oxygen through the blood. Another 400 mg is devoted to cellular proteins that use iron for important cellular processes like storing oxygen in the muscle (myoglobin), performing energy-producing redox reactions (cytochromes FeS cluster proteins) or oxidations (cytochrome enzymes or (cytochromes, enzymes having Fe in the catalytic centre). 3-4 mg circulates through the plasma, bound to transferrin. Some iron in the body is stored. Physiologically, most stored iron is bound by ferritin molecules. The largest amount of ferritin-bound iron is found in cells of the liver hepatocytes, the bone marrow and the spleen. The liver's stores of ferritin are the primary physiologic source of reserve iron in the body. 8

Iron distribution in the human body y

g 2.5 0.15 0.003 1.0 0.0001 0.02 % 68 4 0.1 27 0.004 0.6

hemoglobin myoglobin transferrin ferritin, tissue ferritin, serum enzymes

Iron requirement (if the absorption efficiency is ~10%): male 10 mg/day female 18 mg/day mg/day, Iron sources: meat, liver, leguminoses, fruits
9

Overview of iron metabolism

dietary iron gut absorption plasma transferrin transport

absorption receptors on iron-requiring cells apotransferrin internalization, acidification

synthesis of iron proteins (hemoglobin, myoglobin, (h l bi l bi cytochromes, etc.)

intracellular mobile iron pool

utilization

storage t (mainly in liver) ferritin

No physiologic excretion mechanism! But iron is highly recycled!

hemosiderin

10

Absorption of iron

11

How does the body get its iron? Most of the iron in the body is hoarded and recycled by the y y y reticuloendothelial system (macrophages) which breaks down aged red blood cells. However, people lose a small but steady amount by sweating and by shedding cells of the skin and the mucosal lining of the gastrointestinal tract. The total amount of loss for healthy people in the developed world amounts to an estimated average of 1 mg a day for men, and 1.52 mg a day for women with regular menstrual p periods. People in developing countries with gastrointestinal p p g g parasitic infections often lose more. This steady loss means that people must continue to absorb iron. They do so via a tightly regulated process that under normal circumstances protects against l d h d l i i iron overload.
12

Absorbing iron from the diet Like most mineral nutrients, iron from digested food or supplements is almost entirely absorbed in the duodenum by enterocytes of the duodenal lining. These cells have special molecules that allow them to move i iron i t th b d C ki of f d f ilit t th b kd into the body. Cooking f food facilitates the breakdown of f ligands attached to iron. To be absorbed, dietary iron must be in its ferrous (Fe2+) form Low pH of the stomach reduces iron with the help form. of vitamin C. In addition, a ferric reductase enzyme on the enterocytes' brush border, Dcytb, reduces Fe3+ to Fe2+. A protein called divalent metal transporter 1 (DMT1), which transports all kinds of divalent metals into the body, then transports the iron across the enterocyte's cell membrane and i t th cell. t t ' ll b d into the ll These intestinal lining cells can then either store the iron as ferritin (in which case the iron will leave the body when the cell dies and is sloughed off into feces) or the iron can move it into the body, using a transporter protein called ferroportin. Ferroportin transports Fe2+, but tranferrin carries Fe3+, so iron has to be oxidized by hephaestin on 13 the capillary surface of enterocytes for further transport.

Absorption of iron

Andrews (2005) N. Engl. J. Med., 353, 2508-2509.

Absorption of iron
Stomach Small intestine

Fe2+

Ceruloferroxidases plasmin

low pH, vitamin C

and/or Steap homolog ferrireductases?

Fe3+

HCP1 (heme carrier protein 1)

15

Transport of iron

16

Structure of the transferrin (Tf)

Structure of iron binding site of transferrin

Tf is an 80 kDa serum glycoprotein synthesized mainly by the liver. Tf is bilobal in structure and each lobe contains two domains Both lobes bind iron reversibly and with domains. high affinity. Iron is coordinated by Tyr, Asp and His residues. The binding of iron also needs an anion which is usually carbonate (CO32-). The charge on the anion is balanced by arginine side y ( ) g y g chain. The iron-binding capacity of transferrin is strongly pH-dependent: high-affinity binding occurs at pH 7.4 (Ka ~ 1023 M1), but no binding occurs below pH 4.5. In a healthy individuals only ~30% of Tf binding sites are saturated.

Structure of transferrin receptor 1 (TfR1)

TfR1 is a homodimeric glycoprotein that consists of two 90 kDa subunits linked by disulfide bonds. li k d b di lfid b d
671 aa

The Tf-TfR1 complex occurs in 2:2 stoichiometry stoichiometry. The TfR1-Tf interaction is reversible and is dependent on pH and iron content of transferrin.

S-S
Transmembrane helix

28 aa

TfR2 Role: sensing iron stores. It is

constantly expressed on some iron sensing cells, such as hepatocytes and enterocytes (no IRE in its mRNA). 61 aa 18

How do cells get their own iron? Most of the iron in the body is located on hemoglobin molecules of red blood cells. When red blood cells reach a certain age, they are degraded and engulfed by specialized scavenging macrophages. These cells internalize the iron-containing hemoglobin, degrade it, transport iron via ferroportin molecules into the blood, g , g , p p , which is then transported by the transferrin molecules to the cells expressing transferrin receptors. Most of the iron used for blood cell production comes from this cycle of hemoglobin recycling. All cells use some iron, and must get it from the circulating blood. Since iron is tightly bound to transferrin, cells throughout the body have receptors for transferrin-iron complexes on their surfaces and takes them up by receptor mediated endocytosis. Once inside, the cell transfers the iron to ferritin, the internal iron storage molecule, and recycles of complex of apotransferrin-TrfR back to cell surface where release apotransferrin to blood.

19

Receptor mediated endocytosis of iron-Tf-TfR1 complex iron Tf TfR1

apotransferrin release binding of loaded transferrin to its receptor cell membrane clathrin-coated pits

each cycle takes 2-15 minutes

internalization into coated vesicles

recycling of complex of apotransferrin-TfR1 endosomal pH drop: iron release

intracellular iron pool

20

Storage of iron

21

Structure of ferritin
24-mer of light chain
(L-chains catalyse the formation of iron core

24-mer of heavy chain

(H-chains have ferroxidase activity)

Ferritin is F iti i a water-soluble molecule consisting of 24 subunits th t f t l bl l l i ti f b it that forms a hollow sphere that houses up to 4,500 atoms of iron. Each subunit is one of two isoforms, the heavy (21 kDa) and light (19 kDa) subunits. Ferritin takes up and releases iron from its inner core through hydrophilic channels found in the apoferritin shell. The core contains22 crystal-like Fe(III)-hydroxide-phosphate.

The ratio of heavy-to-light subunits of ferritin varies depending on cell type

Mw (kDa) H24L0 550 pI 4.6
HeLa

cancer
Heart

Kidney

Liver Spleen

H0L24 460 5.7

Nat. Apo

iron overload l d
23

Hemosiderin

Hemosiderin is a water-insoluble iron-protein aggregates present in lysosomes and is a by-product of ferritin degradation through incomplete lysosomal processing It is usually found in liver spleen and bone processing. liver, marrow. Iron stored in hemosiderin is more inaccessible and less 24 effective in producing free radicals than iron stored in ferritin.

Regulation of iron metabolism

at the level of iron uptake!

Main logic of regulation of human iron metabolism Iron is such an essential element of human life, in fact, that humans have no physiologic regulatory mechanism for excreting iron. Human bodies tightly regulate iron absorption and recycling and prevent iron overload solely by regulating iron absorption. Those who cannot regulate absorption well enough get disorders of iron overload ( (haemochromatosis). In these diseases, the toxicity of iron starts ) , y overwhelming the body's ability to bind and store it. Haemochromatosis, is a hereditary disease characterized by excessive absorption of dietary iron resulting in a pathological increase in total body iron stores. Excess iron accumulates in tissues and organs disrupting their normal function. The most susceptible organs include the liver, adrenal glands, the heart and the pancreas; patients can present with cirrhosis, adrenal insufficiency, heart failure or diabetes mellitus. Iron overload may be also the consequence of repeated blood transfusions, or diseases that affect the gastrointestinal tract such as Crohns or celiac disease. Since so much iron is required for hemoglobin, iron deficiency anemia is the first and primary clinical manifestation of i d i li i l if t ti f iron d fi i deficiency. O Oxygen t transport i so t is important to human life that severe anemia harms or kills people by depriving their organs of enough oxygen. Iron-deficient people will suffer or die from organ damage well before cells run out of the iron needed for intracellular processes like electron transport. 26

Main logic of human iron g metabolism regulation

1. humans have no physiologic regulatory mechanism for excreting iron, but we continually loose iron because of loosing enterocytes skin cells sweting enterocytes, or bleeding (enteral infections) 2. human bodies tightly regulate iron absorption and recycling 3. human bodies prevent iron overload solely by regulating iron absorption. If this is damaged absorption (genetically or coupled to other diseases such as coeliakia or Crohn disease) haemochromatosis develops.

27

Summary of human iron metabolism

dietary iron gut absorption absorption Export of iron via ferroportin receptors on iron-requiring cells apotransferrin internalization, acidification plasma transferrin transport

Engulfment of dead cells (erythrocytes) by macrophages synthesis of iron proteins (hemoglobin, myoglobin, (h l bi l bi cytochromes, etc.)

intracellular mobile iron pool

utilization

storage t (mainly in liver)

Loss of iron

ferritin

No physiologic excretion mechanism! But iron is highly recycled!

hemosiderin

28

The body regulates iron levels by regulating absorption of iron in enterocytes y

Factors affecting iron absorption 1. 1 total iron stores 2. the extent to which the bone marrow is producing new red blood cells 3. the concentration of hemoglobin in the blood 4. the oxygen content of the blood 5. inflammation in the body

Regulatory proteins of iron g yp metabolism -HFE -Hepcidin

30

The liver is the central regulator of iron homeostasis. Research over the last decade has confirmed that the liver is the primary site of expression of many of the molecules responsible for the regulation of iron homeostasis. The hereditary hemochromatosis p g y (abnormal accumulation of iron) associated molecules HFE (hefaestin), hepcidin expressed at high levels in the liver. Mouse models of HH, where the genes have been disrupted or mutated all result in hepatic iron overload. Constitutive over-expression of p p p hepcidin (negative regulator) in the liver results in iron deficiency anemia. Liver-specific deletion of HFE in mice recapitulates the phenotype of HH.These studies all suggest a major role for the liver in iron metabolism. Our bodies' rates of iron absorption appear to respond to a variety of interdependent factors, including total iron stores, the extent to which the bone marrow is producing new red blood cells, the concentration of hemoglobin in the blood, and the oxygen content of the blood. We also absorb less iron during times of inflammation. Recent discoveries f f demonstrate that hepcidin regulation of ferroportin is responsible for the syndrome of anemia of chronic disease. The body Th b d regulates i l t iron l levels b regulating each steps of absorption of i l by l ti h t f b ti f iron i in enterocytes. This is achieved within the crypt cells, which sense the availability of iron by taking up iron via both transferrin receptor 1 and 2.The affinity of transferrinbound iron to TfR is affected by a non classical MHC I molecule HFE HFE interacts non-classical molecule, HFE. with TfR1 in such a way that binding of HFE to the TfR enhances its affinity for irontransferrin, resulting in an increase of cellular iron uptake. Depending on the amount of iron, iron following the division of the crypt cells the daughter villus cell responsible for the cells, cell, uptake of the iron, will express the appropriate amount of the Dcytb, DMT1 and ferroportin .
31

Iron influx into enterocyte is determined by the set-point of precursor cells set point
Stomach Small intestine Fe2+
Ceruloferroxidases plasmin

villus cell (enterocyte)

low pH, vitamin C

and/or Steap homolog ferrireductases?

the daughter villus cell cell, responsible for the uptake of the iron from gut, will express the appropriate amount of the DMT1 DMT1, ferroportin proteins.

Fe3+

HCP1 (heme carrier protein 1)

Stomach

Small intestine

Fe2+

crypt cell (precursor cell)

Ceruloferroxidases plasmin

low pH, vitamin C

and/or Steap homolog d/ St h l ferrireductases?

Fe3+

crypt cells, sense th availability of iron t ll the il bilit f i by taking up iron via transferrin receptor 1, 2 helped by the HFE protein.

HCP1 (heme carrier protein 1)

32

Iron influx into enterocyte is determined y by the set-point of precursor cells

In response to iron deficiency anaemia: villus cells produce more Dcytb, DMT1 and ferroportin. In response to iron overload: Villus cell produce less Dcytb, DMT1 and ferroportin.

33

Hepcidin, a circulating peptide hormone, is the master regulator of systemic iron homeostasis, coordinating the use and storage y , g g of iron with iron acquisition. This hormone is primarily produced by hepatocytes in response to iron overload or inflammation. It is i a negative regulator of i ti l t f iron entry i t plasma. H t into l Hepcidin idi functions to reduce serum iron levels by reducing intestinal iron absorption and iron release from macrophages and other cell types and achieves this by binding to the iron exporter ferroportin on the surface of cells and inducing its p g internalisation and degradation. Ferroportin is distributed throughout the body on all cells which store iron. Thus, regulation of f l ti f ferroportin i th b d ' main way of regulating th ti is the body's i f l ti the amount of iron in circulation.
34

Regulatory pathways of hepcidin

35
Chua et al. (2007) Crit. Rev. Clin. Lab. Sci., 44, 413-459.

Hepcidin has antimicrobial properties

high iron level in patients having hemochromatosis makes them more susceptible for microbial infection

inflammation infection i f ti

Macrophage

IL-6 IL 6 Hepatocyte Hepcidin

low iron level

Iron release from enterocytes and macrophages

36

Iron and bacterial protection A proper iron metabolism protects against bacterial infection. If bacteria are to survive, then they must get iron from the environment. Disease-causing bacteria do this in many ways, including releasing iron-binding molecules called siderophores and then reabsorbing them to recover iron, or scavenging iron from hemoglobin and transferrin. The harder they have to work to get iron, the greater a metabolic price they must pay. That means that iron-deprived bacteria reproduce more slowly. So our control of iron levels appears to be an important defense against b t i l i f ti d f i t bacterial infection. P People with i l ith increased amounts of i d t f iron, lik like people with hemochromatosis are more susceptible to bacterial infection. To obtain a more perfect protection during bacterial infection, cytokines (such as IL-6) IL 6) released from the inflammation sites will induce the release of hepcidin sites, hepcidin. (Hepcidin alone is antifungal, and was discovered in urine during a screen for antimicrobial peptides.) Hepcidin functions to reduce serum iron levels, thus decreasing the amount of easily assessable circulating iron for bacterias Although bacterias. this mechanism is an elegant response to short-term bacterial infection, it can cause problems when inflammation goes on for longer. Since the liver produces hepcidin in response to inflammatory cytokines hepcidin levels can increase as cytokines, the result of non-bacterial sources of inflammation, like viral infection, cancer, auto-immune diseases or other chronic diseases. When this occurs, the sequestration of iron appears to be the major cause of the syndrome of anemia of chronic disease, in which not enough iron is available to produce enough 37 hemoglobin-containing red blood cells.

Haemochromatosis: disorders of iron overload

Hemosiderin is deposited allover the body. Haemochromatosis a hereditary disease characterized by excessive absorption of dietary iron resulting in a p pathological increase in total body iron stores.(See models). Excess iron g y ( ) accumulates in tissues and organs disrupting their normal function. The most susceptible organs include the liver, adrenal glands, the heart and the pancreas; patients can present with cirrhosis, adrenal insufficiency, heart failure or diabetes mellitus. (Iron overload may be also the consequence of repeated blood transfusions, or diseases that affect the gastrointestinal tract such as Crohns or celiac disease.) 38

Crypt-programming model of hemochromatosis

39

Liver hepcidin model of hemochromatosis

40

Regulation of iron metabolism at th l t b li t the level of l f cells

What is regulated?
Ferritin concentration Number of tansferrin receptor p
depends on iron amount inside of the cell

1. 1 When iron level is high :

not necessary to uptake more iron, so less transferrin receptor is required to expressed on the cell surface, but more ferritine is required to store excess iron

2, When iron level is low inside the cell:

No need to express storage protein (ferritine), but more receptor is required to take up more iron

The expression of these two protein is regulated paralely an a recyprocal way by the same regulatory protein!

42

In human cells, the best characterized iron-sensing mechanism is the result of translational regulation of mRNA of proteins involved in iron metabolism: transferrin receptors, and for ferritin. When iron level is low inside the cell an iron sensing protein (IRE-BP, a FeS cluster protein) binds to special mRNA sequences of ferritin and transferrin receptor mRNA and thereby inhibiting ferritin translation (synthesis of ferritin protein) but promoting transferrin receptor synthesis (by stabilising its mRNA). When iron level is high iron binds to the iron sensing protein (IRE-BP) the protein changes shape with the result that the it can no longer bind the ferritin and transferrin receptor mRNA, as a consequence the result is just the oposite seen above, so transferrin is readily translated , but no transferrin made. (Interestingly, in iron-bound state the IRE-BP functions as a cytosolic aconitase.) In low-iron conditions IRE-BPs allow the cell to keep producing transferrin receptors And low iron conditions, IRE BPs receptors. more transferrin receptors make it easier for the cell to bring in more iron from transferriniron complexes circulating outside the cell. But as iron binds to more and more IRE-BPs, they change shape and unbind the transferrin receptor mRNA. The transferrin receptor mRNA is rapidly degraded without the IRE-BP attached to it. The cell stops producing transferrin receptors. When the cell has obtained more iron than it can bind up with ferritin or heme molecules, more and more iron will bind to the IRE-BPs. This will initiate ferritin production to store the excess iron iron. (Detailed mechanism: the special mRNA sequences (called iron response elements=IRE) located at different ends of the two mRNAs. If it is located at the 5 end, binding of IRE-BP inhibits translation of th mRNA. If it i l i hibit t l ti f the RNA is located at th 3 end, it protects mRNA f t d t the d t t RNA from degradation leading to more protein synthesis. ) 43

IRE-BP

IRE

IRE

44

SUMMARY

+Fe +F -Fe

45

Utilization of iron

- Heme proteins: hemoglobin, myoglobin, cytochromes, oxidases, peroxidases -I Iron-sulfur cluster proteins: lf l t t i ferredoxin, succinate dehydrogenase, aconitase, etc. - Other iron containing proteins: amino acid hydroxylases (Phe, Tyr, Pro, Lys), acid phosphatase, homogentisinate dioxygenase, ribonucleotide reductase g yg ,

46

HEME / HAEM METABOLISM

Structure of heme
COOH COOH

N Fe N

N N

Fe-protoporphyrin IX
Porphyrins are cyclic compounds that readily bind metal ionsusually Fe2+or Fe3+, and formed by the linkage of four pyrrole rings through methenyl bridges. The most prevalent metalloporphyrin in humans is heme, which consists of one ferrous (F 2+) i f (Fe2+) iron i ion coordinated i th center of h t t di t d in the t f he tetrapyrrole ring of proto l i f t porphyrin IX. Heme is the prosthetic group for hemoglobin, myoglobin, the cytochromes, catalase, 48 nitric oxide synthase, and peroxidase.

Synthesis of heme

49

Tetrapyrrole biosynthetic pathways

(in most bacteria and plants) (5-aminolevulinate) ( (in most eukaryotes) y )

corin ring

porphyrin ring

50

Porphyrins are cyclic compounds that readily bind metal ionsusually Fe2+or Fe3+. The most prevalent metalloporphyrin in humans is heme, which consists of one ferrous (Fe2+) iron ion coordinated in the center of he tetrapyrrole ring of proto porphyrin IX IX. Heme is the prosthetic group for hemoglobin, myoglobin, the cytochromes, catalase, nitric oxide synthase, and peroxidase. These heme proteins are rapidly synthesized and degraded. (For example, 67 g of hemoglobin are synthesized each day to replace heme lost through the normal turnover t rno er of er throc tes Coordinated with the t rno er of heme proteins is the sim ltaneo s erythrocytes. ith turnover heme-proteins simultaneous synthesis and degradation of the associated porphyrins, and recycling of the bound iron ions. Structure of porphyrins Porphyrins are cyclic molecules formed by the linkage of four pyrrolerings through methenyl bridges (F Slide : ). Three structural features of these molecules are relevant to understanding their medical significance. 1. Side chains: Different porphyrins vary in the nature of the side chains that are attached to each p p y y of the four pyrrole rings: Uroporphyrin contains acetate (CH2COO) and propionate(CH2CH2COO) side chains, Coproporphyrin contains methyl(CH3) and propionate groups, Protoporphyrin IX (and heme) contains vinyl (CH=CH2) methyl and propionate groups (CH=CH2), methyl, groups. The methyl and vinyl groups are produced by decarboxylation of acetate and propionate side chains, respectively. 2. Distribution of side chains: The side chains of porphyrins can be ordered around the tetrapyrrole nucleus four tetrap rrole n cle s in fo r different ways, designated b Roman n merals I to IV Onl T pe III a s by numerals IV. Only Type porphyrins, which contain an asymmetric substitution on ring D (see Figure21.2), are physiologically important in humans. (Protoporphyrin IX is a member of the Type III series.) 3. Porphyrinogens: These porphyrin precursors (for example, uro-porphyrinogen) exist in a chemically reduced, colorless form, and serve as intermediates between porphobilinogen and the oxidized, colored protoporphyrins in heme biosynthesis 51

Overview of heme synthesis

mitochondrion
Gly + Suc-CoA
pyridoxal phosphate

HEME
Fe2+

protoporphyrin IX 7 protoporphyrinogen IX 6

-aminolevulinic acid (ALA)

2 Porphobilinogen (PBG) 3 aminomethyl-bilane aminomethyl bilane 4 uroporphyrinogen III 5 coproporphyrinogen III

cytoplasm
The organs mainly involved in heme synthesis are the liver and the bone marrow. 52

Biosynthesis of heme (1)

The major sites of heme biosynthesis are the liver, which synthesizes a number of heme j y y proteins (particularly cytochrome P450 proteins), and the erythrocyte-producing cells of the bone marrow, which are active in hemoglobin synthesis. (Over 85% of all heme synthesis occurs in erythroid tissue.) In the liver, the rate of heme synthesis is highly variable, responding to alterations in the cellular heme pool caused by fluctuating demands for heme proteins In proteins. contrast, heme synthesis in erythroid cells is relatively constant, and is matched to the rate of globin synthesis. The initial reaction and the last three steps in the formation of porphyrins occur in mitochondria, whereas the intermediate steps of the biosynthetic pathway occur in the cytosol. (Slide. ). (Mature red blood cells lack mitochondria and are unable to synthesize heme.) 1. 1 Formation of aminolevulinic acid (ALA): All the carbon and nitrogen atoms of the porphyrin molecule are provided by glycine (a nonessential amino acid) and succinyl coenzyme A (an inter-mediate in the citric acid cycle) that condense to form ALA in a reaction catalyzed by ALA synthase (ALAS) .This reaction requires pyridoxal phosphate (PLP) as a coenzyme, and is the committed and rate-limiting step in porphyrin biosynthesis. (There are two isoforms of ALAS, 1 and 2, each controlled by different mechanisms. Erythroid tissue produces only ALAS2,the gene for which is located on the X-chromosome. Loss of function mutations in ALAS2 result in X linked sideroblastic anemia ) X-linked anemia.) 2. Formation of porphobilinogen: The condensation of two molecules of ALA to form porphobilinogen by Zn-containing ALA dehydratase (porphobilinogen synthase) This enzyme is extremely sensitive to inhibition by heavy metal ions, for example, lead that replace the zinc. This inhibition is, in part, responsible for the elevation in ALA and the anemia seen in lead poisoning. 53

Biosynthesis of heme (2)

3. Formation of uroporphyrinogen: The condensation of four porphobilinogens produces the linear tetrapyrrole, hydroxymethyl-bilane, which is isomerized and cyclized by uroporphyrinogen III. uroporphyrinogen-III synthase to produce the asymmetric uroporphyrinogen III This III. cyclic tetrapyrrole undergoes decarboxylation of its acetate groups (to form methyl), generating coproporphyrinogen III. These reactions occur in the cytosol. 4. Formation of heme: Coproporphyrinogen III enters the mitochondrion, and two propionate side chains are decarboxylated to vinyl groups generating protoporphyrinogen IX, which is oxidized to protoporphyrin IX. Protoporphyrinogen IX oxidase converts the methylene bridges between the pyrrole rings to methenyl bridges (the first colored product).The introduction of iron (as Fe2+) into protoporphyrin IX occurs spontaneously, but the rate is enhanced by ferro-chelatase, an enzyme that, like ALA dehydratase, is inhibited by lead.

54

Overview of heme synthesis

Reaction catalyzed by ALA synthase is the rate-limiting reaction of heme synthesis and it is therefore tightly regulated synthesis, regulated.

ALA d h d t dehydratase

4
Aminomethyl -bilane

Side chains: A: acetyl; P: prppionyl; M. methyl; V: vinyl.

55

Conversion of Uroporphyrins to Coproporphyrins

COO| CH2 |
Acetyl(A)

H+

CO2

CH3 |
Methyl(M)

4x

56

Conversion of Coproporphyrins to Protoporphyrins

COO| CH2 | CH2 |

Propionyl(P)

CO2

CH2 | CH |
2x
Vinyl(V)
57

Steps of Heme Synthesis (7)

Protoporphyrinogen IX oxidase converts the methylene bridges between the pyrrole rings to methenyl bridges. 58

facultative information

Lead poisoning
Catalytic residues of ALA dehydratase are lysines; however reduced cysteines and Zn2+ ion are also needed however, for activity. ALAD is very susceptible to inhibition by heavy metals, especially lead. Increased urinary excretion of its substrate is a good indicator of lead poisoning. Lead poisoning is one of the most common acquired environmental diseases because of the widespread distribution of the metal in Nature and its physical properties which man has utilized for the past 6000 years properties, years. The common symptoms include severe abdominal pain, peripheral neuropathy, constipation, temporary psychiatric disturbance and elevated excretion of haem biosynthetic intermediates. The use of lead by the Romans, not only in water-management schemes but also as artificial sweeteners in the form of lead salts, has been used as an explanation for the lower birth rates and the madness of the Roman upper classes (the Patricians), and has been linked to the decline of the Roman Empire. Another disastrous episode that has been attributed to lead poisoning, at least in part, was the ill-fated Franklin expedition to the Arctic, in search of the Northwest passage, which set out from England in May 1845. Sir John Franklin led the voyage; he was in charge of two ships, Erebus and Terror, and 128 crewmen. The expedition was extremely well organizedFranklin had ample provisions, fresh fruit juice (to prevent scurvy), modern organized Franklin equipment and well-trained officersbut no members of the crew ever returned. It is known that the two ships reached Beechey Island and sheltered in the harbour during the winter of 18451846. During this period, three crew members died and were buried in the permafrost. The evidence suggests that the remainder of the expedition became disorientated and disorganized, and vanished into the icy wasteland during the subsequent months. An analysis of the frozen and perfectly preserved bodies of the three crew members buried on Beechey Island revealed that they had very high levels of lead in their bodies. A similar study of the remains of some of the crew found on King William Island also supports this claim. Thus it has been suggested that the c e o t e e ped t o su e ed o ead po so g, crew of the expedition suffered from lead poisoning, because lead containers were used to store water and ead co ta e s e e sto e ate a d food, and that the subsequent symptoms of muscle weakness and mental derangement contributed to the demise of the expedition. 59
Warren et al. (1998) TIBS, 23, 217-221.

facultative information

Names of Porphyrins
They consist of a word and a number e g uroporphyrin III The word denotes the kinds of number, e.g., III. substituents found on the ring, and the number denotes how they are arranged. WORDS: uroporphyrin, coproporphyrin, protoporphyrin
AP MP MPV

NUMBERS: I, II, III, IV I II III In series I the substituents repeat in a regular manner: AP AP AP AP. Series II does not occur in natural systems. In series III the order of substituents in ring IV is reversed: AP AP AP PA. g Series IV does not occur in natural systems. Porphyrin vs Porphyrinogen Porphyrinogens are more reduced than the corresponding porphyrins porphyrins. The porphyrins contain a system of conjugated double bonds all around the tetrapyrrole ring, 60 which makes the porphyrins more stable than the corresponding porphyrinogens.

Regulation of Heme Synthesis

synthesis of new enzyme

HEME
Fe2+

cytoplasm

ALA synthase

mitochondrion it h d i
Gly + Suc-CoA
pyridoxal phosphate

1 ALA synthase

protoporphyrin IX 7 protoporphyrinogen IX 6

-aminolevulinic acid (ALA) i l li i id

2 porphobilinogen 3 aminomethyl-bilane 4 uroporphyrinogen III 5 coproporphyrinogen III

61

Regulation of Heme and Globin Synthesis

- Substrate availability: iron (II) must be available for ferrochelatase. - Feedback regulation: heme is a feedback inhibitor of ALA synthase. - Subcellular localization: ALA synthase is in the mitochondria, where th substrate, succinyl C A i produced. h the b t t i l CoA, is d d ALA synthase is synthesized in the cytoplasm, its transport across the mitochondrial membrane may be regulated. - In erythropoietic cells, heme synthesis is coordinated with globin synthesis. If heme is available, globin synthesis proceeds. If heme is absent: - Effects of drugs: barbiturates and certain steroids can increase heme synthesis via increased production of the rate-limiting enzyme ALA synthase enzyme, synthase.

62

Porphyrias
Defects in heme synthesis The porphyrias are classified depending on whether the enzyme deficiency occurs y In the erythropoietic cells of the bone marrow: Erythropoietic In the liver: Hepatic Either type may be hereditary or acquired. The symptoms are caused by accumulation of intermediates and deficiency of heme. Accumulated intermediares are converted by nonenzymatic (light, oxidative effects) steps f id ti ff t ) t from porphyrinogens t unuseful/ porphyrins h i to f l/ h i which makes photosensitivity. Porphyria refers to the purple color caused by pigment like por Porphyria pigment-like porphyrins in the urine.

63

Defects in heme synthesis

Pb poisoning

2
Pb

Porpyrias of erithropoietic origin

1: erithropoietic porphyria 2: hereditary protoporphyria 3: acute intermittent porphyria 4: porphyria cutanea tarda 5: hereditary coproporphyria 6: variegate porphyria

Porphyrias of liver origin

64

Porphyrias
Sympthomes: 1. individuals with an enzyme defect prior to the synthesis of the tetrapyrroles manifest abdominal and neuro -psychiatric signs signs, 2. enzyme defects leading to the accumulation of tetrapyrrole intermediates show photosensitivitythat is, their skin is itches and burns (pruritus) when exposed to visible light. (Photosenstivity is a result of the oxidation of colorless porphyrinogens to colored porphyrins, which are photosensitizing molecules that are thought to participate in the formation of superoxide radicals from oxygen. These reactive oxygen species can oxidatively damage membranes, membranes and cause the release of destructive enzymes from lysosomes.)

red urine,

injured skin

65

Acute intermittent porphyria

ALA synthase Glycine + Succinyl-CoA Succinyl CoA ALA ALA PBG deaminase PBG // ... PBG no feedback inhibition! // Heme Heme Hemoproteins

Porfobilinogen (PBG) deaminase deficiency on one chromosome The remaining chromosome. activity is sufficient to produce heme for erythropoiesis. In the liver, however, if heme is utilised or degraded by an elevated rate (e.g. certain drugs, hormones or ethanol are metabolised by cytochrome P450 containing enzymes, they induce the level of this heme y y g y , y containing enzyme) the decrease in the levels of heme induces ALA synthase. Under these conditions the elevated levels of PBG cannot be further converted by PBG deaminase. Both PBG (red urine) and ALA (neurotoxicity) are accumulated. Symtomps appear always after the appearance of the triggering molecule in the form of acute abdomen syndrom, neurological abnormalities. Can be treated by infusion of high concentration of heme. Barbiturates must be avoided beacuse they increase the level of ALA synthase. y
66

Summary of heme synthesis

- It occurs in virtually all tissues but the highest rate is found in the liver and b li d bone marrow. - The first and the last three enzymes are located in the mitochondria. The middle 4 enzymes are located in the cytosol. - Heme is synthetized from 8 glycine and 8 succinyl-CoA molecules. y gy y - During synthesis the side chain modifications occur on the colorless porphirinogens. porphirinogens - The last step oxidizes it to porphyrin (methylen to methene bridges) which has color color. - Porphyrins are produced by nonenzymatic (light, oxidative effects) steps f t from porphyrinogens i porphyrias. h i in h i
67

Degradation of heme

68

Degradation of heme

69

Recycled!

A HEME degradation
slpeen, macrophages

Heme oxigenase

Biliverdin

Biliverdin reductase
UDP glkuronil transzferz

LIVER

Bilirubin BLOOD
(albumin)

INTESTINE BILE

Bilirubin Bacterial flora dconjugation, redcution Saturation of methenyl

Reduciton ,viniletil conversion vinil etil

KIDNEY

70 feces Stercobilin urine Urobilin

Bilirubin
The high lipid solublity of bilirubin dictates - that it must be transported in the blood by a carrier (serum albumin) - that it is soluble in the lipid bilayers of cell membranes - that it must be conjugated to a water-soluble substance for excretion Bilirubin diglucuronide is excreted in the bile. It is subject to subsequent transformations to other species by the intestinal fl t f ti t th i b th i t ti l flora. The clinical determination of plasma bilirubin distinguishes between conjugated (direct) and unconjugated (indirect) bilirubin bilirubin. - Direct and indirect bilirubin values are used in the differential diagnosis of hyperbilirubinemia.

71

Jaundice (also called icterus) refers to the yellow color of skin, nailbeds, and sclerae (whites of the eyes) caused by deposition of bilirubin, secondary to increased bilirubin levels in the blood ( yp (hyperbili rubinemia). Although not a disease, jaundice is usually a symptom of an underlying ) g j y y p y g disorder. Types of jaundice: Jaundice can be classified into three major forms described below. However, in clinical practice, jaundice is often more complex than indicated in this simple classification. For example, example the accumulation of bilirubin may be a result of defects at more than one step in its metabolism. a. Hemolytic jaundice: The liver has the capacity to conjugate and excrete over 3,000 mg of bilirubin per day, whereas the normal production of bilirubin is only 300 mg/day. This excess capacity allows the liver to respond to increased heme degradation with a corresponding p y p g p g increase in conjugation and secretion of bilirubin-diglucuronide. However, massive lysis of red blood cells (for example, in patients with sickle cell anemia, pyruvatekinase or glucose 6-phosphate dehydrogenase deficiency) may produce bilirubin faster than it can be conjugated. Unconjugated bilirubin levels in the blood become elevated causing jaundice (More conjugated bilirubin is elevated, excreted into the bile, the amount of urobilinogen entering the enterohepatic circulation is increased, and urinary urobilinogen is increased.] b. Hepatocellular jaundice: Damage to liver cells (for example,in patients with cirrhosis or hepatitis) can cause unconjugated bilirubin levels in the blood to increase as a result of decreased conjugation. Urobilinogen is increased in the urine because hepatic damage decreases the enterohepatic circulation of this compound, allowing more to enter the blood, from which it is filtered into the urine. The urine thus darkens, whereas stools may be a pale clay color Plasma levels of AST and ALT(special liver enzymes) are elevated [Note: pale, color. elevated. If conjugated bilirubin is not efficiently secreted from the liver into bile (intra-hepatic cholestasis), it can diffuse (leak) into the blood, causing a conjugated hyperbilirubinemia.] The similar thing hapens in case of neonatal jaundice. c. Obstructive jaundice: In this instance, jaundice is not caused by overproduction of bilirubin or decreased conjugation, but instead results from obstruction of the bile duct (extrahepatic 72 cholestasis).

Hyperbilirubinemia / Jaundice

Normal

Hemolytic jaundice y j
73

Hyperbilirubinemia / Jaundice

Normal

Physiological (neonatal) jaundice

In neonates, benign jaundice tends to develop because of two factors: - the breakdown of fetal hemoglobin as it is replaced with adult hemoglobin - immature hepatic metabolic pathways which are unable to conjugate and so excrete bilirubin as quickly as an adult. Infants with neonatal jaundice are treated with colored light called phototherapy. f Phototherapy works through a process of isomerization that changes the bilirubin into water-soluble isomers 74 that can be passed without getting stuck in the liver.
Wikipedia

Hyperbilirubinemia / Jaundice

Normal

Biliary obstruction

75

HEMOGLOBIN

76

Functions of Hemoglobin
Lung Circulation Tissues

inhaled

O2

Hb.4O2

O2

respiratory chain

exhaled

TCA cycle
CO2 + H2O

CO2 + H2O

carbonic anhydrase
H2CO3 H2CO3

carbonic anhydrase

H+

+ HCO3

Hb.2H Hb 2H+ Hb.carbamate

H+ + HCO377

Quaternary structure of hemoglobin

1
Hemoglobin Quaternary structure: 4 subunits! Four polypeptide chains Four Four heme, four Fe2+, four O2 The 4 monomer are kept together by secondary bonds: y Salt bridges Hydrogen bonds

78

HgA1: 22

79

Structure of one subunit of hemoglobin

Tertiary structure of globin chain comprises 8 -helices: helices:
helix A: Ser3- Gly18 helix B: His20-Ser35 helix C: Phe36-Tyr42 helix D: His50-Gly51 helix E: Ser52-Ala71 helix F: Leu80-Ala88 helix G: Asp94-His112 helix H: Thr118-Ser138 (Name of the loop between two helices is composed from names of the two helices: eg. AB, CD)

The hem group is found in the apolar pocket of the globin molecule with its polar groups facing the surface. 80

Structure of heme
COOH COOH

N Fe N

N N

Heme is the prosthetic group for hemoglobin (myoglobin) Heme consists of one ferrous (Fe2+) iron ion coordinated in the center of he tetrapyrrole ring of proto porphyrin IX.

Fe-protoporphyrin IX

81

Tertiary structure of hemoglobin

Haemoglobin is a globular very tightly stuffed compaque molecule. Hydrophobe inside, y p , hydriphyl side chains outside. Heme is inside of the hydrophobe p pocket. Isolated fee heme unable to keep Fe in 2+ state, only pocked inside the globin chain. If Fe is oxidized to Fe3+ (ferri) ( ) (methemoglobin) it cannot bind O2. Iron has 6 coordinative (covalent ) bindings: 1-4.: 1 4 : 4 Nitrogene of porphyrine 5.: His-F8 of globin (proximal His) This makes the bond between globin and Heme 6.: O2 His-E7 helps to bound O2 (distal His)
(no O2 is show here)

proximal His

oxygen binding site g

distal His
82

Polimorfism of globins
I. II. III.

I.Embrionyc haemoglobins Hb Gower 1 Hb Gower 2 Hb P tl d Portland II.Foetal Haemoglobins Hb F III.Adult haemoglobins HbA1 HbA2
(98%) (2%)

22 22 22

22

2 2 22
83

Expression of hemoglobin genes during development is related to the changing oxygen uptake

Comparison of Mb and Hb

Myoglobin One polypeptide chain p yp p One heme, one Fe, one O2 Tertiary structure only No cooperativity O2 storage in muscle

Hemoglobin Four polypeptide chains p yp p Four heme, four Fe, four O2 Tertiary and quaternary structure Cooperativity Regulated affinity to O2 binding 84 O2 transport in RBC

Oxygen-binding to Hb yg g
100 80 Oxygen-binding curve for Hb

- sigmoidal / cooperative - low affinity in the veins - high affinity in the arteries g y - p50 25 mmHg

% saturat tion

60 40 20 0
venous pressure arterial pressure

20

40

60

80

100

pO 2 (mmHg)
85

O2-binding to Mb g
100 80 Oxygen-binding curve for Hb
Mb

Hb

- hyperbolic / non-cooperative - high affinity for O2, higher than that of Hb g - p50 1 mmHg

% saturat tion

60 40 20 0

20

40

60

80

100

pO 2 (mmHg)
86

O2-binding causes conformational changes

87

The quaternary structure changes q y g

In deoxyHb iron is out of the plane of the heme. Following oxigen binding iron moves g g g into the plane. The movement of iron is followed by the movement of the protein chain attached attached. Upon oxygen binding: subunit salt bridges break 11 twists relative to 22 heme-heme distance reduces central cavity constricts l i i

In short, the deoxy state relaxes and switches to the oxy state. These changes transmits the structural changes to the other g heme groups and INCREASES their O2 binding. This is cooperativity.
88

Cooperativity
100 80 Oxygen-binding curve for Hb

% sa aturation

60 40 20 0

20

40
2

60

80

100

pO (mmHg)

In other words the oxygen binding to the next subunits requires less energy because part of the salt bridges are already broken. That is why the affinity becomes larger. This explains the sigmoid saturation curve. 89

Regulation of O2 binding affinity

O2 affinity is decreased by:
1. 2,3BPG (2,3-bisphospho-glycerate) -produced by the shunt of the glycolysis in RBC -releases O2 2. Low pH- Bohr effect, -metabolically active tissues (CO2 and H+) metabolically -releases O2 High temperature -metabolycally active warm tissues Aminoacid sequence A i id -Foetal Hb binds O2 with more affinity than adult Hb

3. 3 4. 4

90

Control: 1. 2,3-bisphospho-glycerate
2,3 BPG binds to the positively charged amino acids in the pocket between the two beta chains. -helps the relase of oxygen at tissues -reduces O2 affinity

At high altitude the level of BPG increases facilitating the release of oxygen at tissues. At l low external oxygen more 2 3 BPG bi d to the t l 2,3 binds t th increased amount of deoxiHb, 2,3BPG will not inhibit its own production (BPG mutase). Increased BPG synthesis.

91

Glucose Glucose-6-P

Synthesis of 2,3-BPG y ,
Biphosphoglycerate mutase

ADP ATP

1,3-biphosphoglycerate 1 3 biphosphoglycerate 2,3-biphosphoglycerate 3-phosphoglycerate Phosphenolpyruvate Pyruvate y Lactate deoxyhemoglobin

Deoxyhemoglobin + 2 3-BPG 2,3-BPG

At high altitude the level of BPG increases facilitating the g g release of oxygen at tissues. At lowexternal oxygen more 2,3 BPG binds to the increased amount of deoxiHgb, 2,3BPG will y not inhibit BPG mutase. Increased BPG synthesis.
92

Control: 2. The Bohr effect

Metabolically active tissues are rich on CO2 and H+. At such sites Hb releases O2 and picks up CO2 and H+. Why?
R NH 3+ N-terminus of
N NH H+ O His Hi 146 of f Asp A 94 of f O- + HN NH

CO 2

R N CO 2H

These additional charges form additional salt bridges to further g g cross-link the Hb quaternary structure and stabilize the tense deoxy state. Hence, they lead to the release of O2.

93

Control: 2. The Bohr effect

low level of CO2

high level of CO2

Both increased H+ and CO2 concentrations decrease the affinity of Hb for O2

94

Control: 3. Temperature

Hb is a temperature controller. O2 binding to Hb is (usually) exothermic; oxygen release from Hb is endotermic, that is, heat is given out. This also means that when oxyHb arrives at muscle, heat is required to liberate O2. Whilst this isnt generally a problem to humans, it is for animals from colder environments where heat is more precious Hbs in these animals have evolved to reduce the heat precious. Hb s needed to free oxygen. At the other extreme, in the heavily worked flight muscles of some birds, efficient heat loss is essential to avoid overheating. Here O2 release requires 3 times as much heat as it does in man.

95

Control: 4. Amino acid sequence q

HbA: 22 HbF: 22 One of the changes in the chain vs the h i is Hi 143S th chain i His143Ser, which li i hi h lies in the central cavity. This change lowers the affinity of deoxyHbF for BPG relative to deoxyHbA This increases the affinity of HbF for O2.
96

Abnormal hemoglobins
Point mutations in the core region

Hemoglobin M
Change E7 or F8 His to Tyr, therefore Fe2+ oxydizes to Fe3+, therefore it cannot bind O2.
97

Abnormal hemoglobins
Mutations at subunit interfaces

Sickle cell anemia

Hemoglobin S: Glu6Val in chains 98
Wikipedia

Abnormal hemoglobins
Mutations at subunit interfaces

mutation

Altered surface of deoxyHbS causes polymerization 99

Wikipedia

Abnormal hemoglobins
Thalassemias
Missing one of the Hb chain Since Hgb genes have several copies chain. the severity of the disease might vary.

Glucosylated Hb (HbA1c)
Glucose is spontaneously covalently bound to Hb. % of Hb glucosylated depends on blood sugar levels. Significance in the early diagnosis of diabetes mellitus.

100

CONTENTS
I.
1. 2. 2 3. 4.

IRON METABOLISM
Introduction Absorption of iron Transport of iron and Storage of iron g p Regulation of iron metabolism: hepcidin

II.
1. 2.

HEME METABOLISM
Biosynthesis of heme, Porphyrias Degradation of heme, Jaundice

III. HEMOGLOBIN, MYOGLOBIN

1. 2. 3. 3 4. Structure of hemoglobin Polymorphism of globins O2 binding and its regulation by 2 3BPG pH, CO2, temperature 2,3BPG, pH CO2 Abnormal hemoglobins: Sicle cell anemia, MetHb, HbA1c

101

Exam essay q y questions

1. 1 Overwiev of iron metabolism absorption of iron transport and metabolism, iron, storage of iron. Regulation of iron uptake at body and cellular level. Heme synthesis, Porpyrias. Heme breakdown. Jaundice. Hemoglobin: structure and function, Regulation of O2 binding .Globin polymorphysm and abnormalities.

2. 3. 4.

102

Example for Simple questions

Give a short definition to
1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. Ferritin Transferrin Transferrin receptor Hepcidin Ferroportin DMT1 Hemocromatosis Porpyrins Heme ALA synthase Ferrochelatase Porhyrias Hemoxigenase UDP glucuronyl transferase Bilirubin Hemoglobin Myoglobin Cooperativity p y Bohr effect Sicle cell anemia HgA1c

Answer the questions shortly

1. 2. 3. 4. 5. 5 6. 7. 8. 9. List Heme containing proteins of human body! What is the mechanism of iron uptake to cells? What is effects of hepcidine? List 3-5 intermediates of heme synthesis! List 3 5 intermediates of heme breakdown! 3-5 Classification of porphyrias? Types of jaundice and short explanation to them! List the factors which affect O2 binding of Hg! What is the composition of adult and foetal Hg? g

103