Hemoglobin and Iron Metabolism (Video Notes)

Vocabulary-style flashcards covering Hb structure, synthesis, regulation, oxygen binding, disease-related variants, and iron metabolism concepts from the video notes.

Heme

Protoporphyrin IX bound to iron (Fe); can reversibly bind one O2 molecule; gives blood its red color.

Hemoglobin (Hb)

Protein in RBCs composed of heme plus globin; accounts for about 95% of the RBC cytoplasm; Hb concentration is ~34 g/dL and contributes to RBC viscosity and deformability.

Globin

Protein portion of Hb consisting of four polypeptide chains (two pairs); each chain ~141–146 aa (alpha - theta); organized into eight helices (A–H) and seven nonhelical segments.

2 alpha chains

2 non alpha chains

Complete Hemoglobin Molecule

Four globin chains form a pocket for four heme groups; each heme carries one O2, allowing Hb to carry up to four O2 molecules.

4 global, 4 heme+ 4 irons forms →

oxygen

Heme Insertion Site in Hb

Heme is inserted at histidine residues on the E7 and F8 helices of the globin chains.

4 Heme + 4 Globin =

1 Hemogloblin

Heme Synthesis Location

Occurs in bone marrow erythrocyte precursors, in mitochondria and cytoplasm; part of Hb biosynthesis.

Heme Synthesis happens in erthrocyte precursor stages _____ to _______

pronormoblast to polychromatic erythrocyte (retic)

Globin Synthesis Location

Protein synthesis from mRNA in the cytoplasm of erythroid precursors.

Synthesis of Protoporphyrins (Heme) Location

Begins in the mitochondria with the formation of delta-aminolevulinic acid (d-ALA) from glycine and succinyl-CoA

d-ALA Synthesis

is formed from glycine and succinyl-CoA in mitochondria; catalyzed by d-ALA synthase, the rate-limiting step of heme biosynthesis.

d-ALA Synthase

Enzyme that catalyzes the formation of d-ALA; activity is inhibited by vitamin B6 (pyridoxine) deficiency.

Porphyrinogens

Unstable, inactive forms of porphyrins that are intermediates in porphyrin/heme biosynthesis.

Porphyrias

Inherited defects in heme synthesis leading to accumulation of porphyrin precursors.

Iron Delivery to Erythroid Cells

Ferric (Fe3+) iron is transported in plasma by transferrin and is reduced to ferrous (Fe2+) iron inside cells for incorporation into heme.

True or False: Only ferrous iron can be incorporated into heme with ferrochelatase.

True

Ferritin

Storage form of iron; keeps iron in a readily available form; stores iron in liver and other tissues; increased in iron overload, decreased in deficiency.

Transferrin

Plasma protein that binds Fe3+ and transports iron to tissues, including RBC precursors.

Genes for Globlin Biosynthesis for chromosome 16

alpha and zeta

Genes for Globlin Biosynthesis for chromosome 11

beta, gamma delta, epsilon

Hemogloblin A

The main form of hemoglobin in adult humans, composed of two alpha and two beta chains. It is responsible for oxygen transport in the blood.

Hemoglobin A₂

A minor form of hemoglobin in adults, comprising two alpha and two delta chains, constituting about <3.5% of total hemoglobin.

Hemoglobin F

The primary hemoglobin found in the fetus, consisting of two alpha and two gamma chains, it has a higher affinity for oxygen than adult hemoglobin, constituting about 1-2% of total hemoglobin.

Embryology through childhood: week 0-10

epsilon and zeta are in egg yolk, but decrease

Embryology through childhood: week 10-week 0 at birth

during hepatic stage and medullary stage, alpha and gamma starts to increas by a lot, while beta increases slowly

Embryology through childhood: week 0 at birth-week 10 at birth

alpha stays consistent, while beta increases more at week 10. Delta then starts to increase slowly. At week 10 there is the beta-gamma switch

Embryology through childhood: week 10-week 0 at birth

alpha stays consistent, while beta increases more after week 10, predominating over gamma. Delta still increases slowly.

Globlin chains of Intrauterine (IU) in Gower 1

zeta + epsilon

Globlin chains of Intrauterine (IU) in Gower 2

alpha + epsilon

Globlin chains of Intrauterine (IU) in Portland

zeta + gamma

Embryonic/Perinatal Hb Forms

Intrauterine Hb includes Gower and Portland types; HbF predominates at birth; HbA becomes predominant in infancy.

Heme Regulation

Heme inhibits ALA synthase, ◦ Increase heme → decrease amount of ALA synthase

Globin Regulation

• Transcription = Promoter + Kruppel-like factor 1 + other transcription factors + locus control region

• Translation = inhibited by lack of heme; increased in presence of heme

Hemoglobin Regulation

◦ Normally stimulated by tissue hypoxia

◦ Increased EPO release

Hemoglobin Functions in High Affinity

Bind oxygen readily in the lungs

Hemoglobin Functions in Low Affinity

Unload oxygen in the tissues

Affinity is dependent upon ______

partial pressure of oxygen (pO2)

Higher Partial pressure of oxygen →

higher affinity

Lower Partial pressure of oxygen →

lower affinity

Bohr Effect

Oxygen affinity of Hb is modulated by pH; higher pH (alkalosis) increases affinity (left shift), lower pH (acidosis) decreases affinity (right shift).

Right Shift in Oxygen Dissociation Curve

increase of H+ ions, lower blood pH, higher 2,3 BPG, higher pCO₂, higher temperature, lower oxygen affinity → decrease in oxygen affinity

Left Shift in Oxygen Dissociation Curve

decrease of H+ ions, higher blood pH, lower 2,3 BPG, lower pCO₂, lower temperature, higher oxygen affinity→increase in oxygen saturation

Oxygen Dissociation Curve

Plot of Hb oxygen saturation versus pO2; indicates how readily Hb binds/releases O2. Shifts to the left/right is dependent on blood pH (Bohr Effect)

P50

Partial pressure of O2 at which Hb is 50% saturated; normally ~27 mm Hg; lower P50 means higher affinity, higher P50 means lower affinity. ph 7.4

Deoxyhemoglobin

specific spatial configuration that resembles an egg with a central cavity; has 2,3-DPG keeping the structure close together

Oxyhemoglobin

changes overall configuration which facilitates additional binding of oxygen. has no 2,3-DPG

Increase of CO₂ during transport of CO₂ across membrane

Co₂ diffuses across the membrane into red blood cells, promoting the release of oxygen from hemoglobin and enhancing the production of bicarbonate.

Bohr effect in transport of CO₂

has a right shift in the oxygen-hemoglobin dissociation curve, meaning that increased levels of CO2 lower the affinity of hemoglobin for CO₂, facilitating oxygen release to tissues.

What does chloride do in transport of oxygen?

Chloride shifts into red blood cells to balance the negative charge as bicarbonate ions diffuse out, facilitating the transport of oxygen and influencing hemoglobin function.

Globin Gene Clusters

Alpha-like globin genes located on chromosome 16 (e.g., zeta, alpha); beta-like globin genes on chromosome 11 (e.g., epsilon, gamma, delta, beta).

CO2 Transport

CO2 is transported in plasma and as bicarbonate; carbonic anhydrase catalyzes CO2 + H2O to H2CO3, then HCO3-

Methemoglobin

Hb with Fe3+ (ferric) that cannot bind O2 with spectrometry detection at 630 nm; Toxic levels: 1.5g%

Why is methemoglobin a problem?

Fe3+ cannot bind to oxygen → left shift in the oxygen dissociation curve, leading to reduced oxygen delivery to tissues → cyanosis

What color the blood would be for methemoglobin?

Dark chocolate brown

Can methemoglobin be corrected?

yes through Oxygen inhalation and Strong reducing substances

Sulfhemoglobin

Hb modified by irreversible oxidation from certain drugs/chemicals (e.g., sulfonamides); spectrometry detected at 630 nm’ toxic level: 0.5%

Why is sulfhemoglobin a problem

causes a left shift in the oxygen-hemoglobin dissociation curve → cyanosis

Can sulfhemoglobin be corrected?

No, it is irreversible.

Carboxyhemoglobin

heme iron bound to carbon monoxide (CO); spectrometry detected around 540 nm; Toxic level: 20-30% (cherry red color)

Reference Ranges for Carboxyhemoglobin

<3% (nonsmokers); 4-15% (smokers)

Why is carboxyhemoglobin a problem?

CO has an affinity 240x greater than oxygen. CO binds Hgb and prevents oxygen from binding thus leading to incredible cyanosis and massive tissue necrosis

Can carboxyhemoglobin be corrected?

Yes, it can be corrected with oxygen therapy or by removing the source of carbon monoxide.

Dietary Iron Sources

Heme iron from animal products (e.g., Hb, myoglobin) and nonheme iron from legumes and leafy vegetables; heme iron often better absorbed.

Average diet for dietary iron

Average diet: 10-20 mg/day

◦ Actual absorption: 1-2 mg/day

◦ Sufficient for most men, but others may need more

Iron is absorbed in two forms:

heme and nonheme iron.

Prussian Blue (Iron Stain)

Histochemical stain used to detect iron in tissues; gold standard for assessing iron stores in bone marrow.

Ring Sideroblasts

Nucleated erythroblasts with iron-laden mitochondria forming ringed structures around the nucleus, seen with Prussian blue staining.

Serum Ferritin

Storage form of iron in macrophages and secreted by macrophages and can be measured in immunoassay; reference rangeL 40-400 ng/mL

Clinical Significance in Low levels of Ferritin

iron deficiency anemia

Clinical Significance of Acute phase protein in Ferritin

hard to determine deficiency

Clinical Significance in High levels of Ferritin

organ damage, inflammation, pregnancy

TIBC (Total Iron Binding Capacity)

Total available iron-binding sites on transferrin; reflects transferrin concentration; normal range ~250–400 µg/dL; can be used with serum iron to calculate transferrin saturation.

In a normal, healthy

individual, iron occupies _____ of the binding sites on transferrin

20-50%

Serum Iron

Amount of ferric iron bound to transferrin in serum = availability of transport iron; typical reference range 50–160 µg/dL; analyzed in Chromogen-Spectrophotometric Methods

Special considerations for serum iron

◦ Does not include iron in hemoglobin.

◦ Diurnal variation: highest in the morning

◦ Limited utility on its own

% Transferrin Saturation

% of transferrin binding sites that are iron-bound

Calculation of % Transferrin Saturation

[Serum Iron/TIBC] x 100

◦ TIBC = total # of sites for Fe binding

◦ SI = # bound with Fe

Why is % Transferrin Saturation a better diagnostic indicator for disease?

It establishes a relationship

between iron and transferrin.

◦ High in Fe overload; low in Fe deficiency

Distribution of Iron

Transferrin bound ot iron will give to iron in:

75% hemoglobin/erthropoeisis

10-20% in ferritin when it stores in liver and heart

5-15% for myoglobin and other enzymes.

Iron Absorption

Dietary iron (ferric state 3+) goes through stomach converting to ferrous iron (2+). It is then absorbed in the duodenum and jejunum of the small intestine. From there, it is converted to ferric state (3+) to ferritin or transported by transferrin in the bloodstream. It can also stay as ferrous iron (2+) to go to RBC precursors to make hemoglobin.

Enterocyte Iron Handling

In the gut, Fe3+ from the diet is transferred into entereocyte to plasma. It is then reduced and transported by DMT1; ferroportin exports Fe2+ into blood; hepcidin regulates this export.

Iron Route in Body after iron is in transferrin

Blood carries iron in transferrin. It goes into bone marrow where it is utilized for hemoglobin synthesis in red blood cells, and excess iron is stored in the liver and spleen as ferritin.

Divalent Metal Transporter 1 (DMT1)

A transporter protein that facilitates the uptake of ferrous iron (Fe2+) and other divalent metals into enterocytes from the intestinal lumen.

With excess Fe, liver makes ____ that binds to ferroportin preventing Fe+2 binding

hepcidin

Hephaestin

in enterocyte membrane oxidizes Fe+2 to Fe+3

In blood Fe+3 (x2) binds to apotransferrin→ ____

transferrin

Transferrin Receptor (TfR1)

Cell-surface receptor that binds transferrin and mediates iron uptake by endocytosis into cells.

DMT1 _______ out of endosome into cytoplasm

moves Fe

Fe+3 binds to apoferritin =

ferritin

How many Fe3+ can 1 ferritin bind?

400

_______ and _______ store most Fe+3

macrophages, hepatocytes

Apoferritin

protein shell sans iron

Hemosiderin

amalgam of degraded iron (micelle form) and a piece of the ferritin shell

Two forms of Recycling Iron

Extravascular hemolysis

Intravascular hemolysis

Extravascular hemolysis

Macrophage ingests RBC, degrades Hgb releasing Fe which binds to ferritin

Macrophages have ferroportin to export Fe to other cells

Macrophages in the spleen and liver recover iron from hemoglobin and return it to circulation via ferroportin.

Intravascular hemolysis

◦ Haptoglobin/hemopexin in blood bind to Hgb/heme released from lysed

RBCs

◦ Hepatocytes have ferroportin too

Practical Iron Deficiency Definition (WHO)

Iron deficiency defined by ferritin <15 µg/L or transferrin saturation <16% or Hb rise after 2 months of iron therapy.

Acute-Phase Considerations in Iron Studies

Ferritin can be elevated with inflammation; timing and CRP should be considered when interpreting iron studies.

Iron Studies Laboratory Assessment

Serum Iron (SI)

Serum Total Iron Binding Capacity (TIBC)

Ferritin

Percent Transferrin Saturation

Prussian blue staining