Immunoglobulins and Hemoglobins

• hemoglobin is the most abundant blood protein

  • 14-16 g/dL
• immunoglobulin classes

  • IgG - soluble, relatively simple structure, most abundant
  • IgA - mucosal surfaces, skin, salivary
  • IgM - first in immune response
  • IgD
  • IgE - involved in allergies
  • concentration decreases down the list
• immunoglobulins contain heavy chains (γ, α, μ, δ, ε) and light chains (κ or λ)

  • IgG has one light and two heavy
  • disulfide bonds hold heavy to heavy and heavy to light
  • antigen binding site between heavy and light chains
• the immunoglobulin fold - a type of secondary structure

  • CDR = complementary determining region
  • heavy/light chains come from different genes but have similar structures
  • 12 genes code for millions of antibodies
  • b cells making antibodies are not identical, thus make different antibodies
  • diversity happens in CDRs
  • in beta sheets
  • mechanisms to diversify genes at CDRs (where antigen binds)
  • in both heavy and light chains
• the antigen binding site

  • 6 loops provide for most of the sites for antigen binding
  • called hypervariable regions or CDRs
• Hemoglobin (Hb) vs Myoglobin (Mb)

  • Hemoglobin
  • circulating form
  • binds O2 at high pH
  • releases O2 at low pH
  • carries CO2 and H+ back to lungs
  • adapts to conditions of higher or lower O2 in atmosphere
  • Myoglobin
  • Mb structure found before Hb structure
  • facilitates O2 diffusion (and storage) in tissues
  • binds to O2 more tightly than hemoglobin
  • pH change does not change its affinity for O2
  • rests in muscles before O2 is used for diffusion
• heme


  makes blood red, holds iron in blood
• at low pH, hemoglobin shifts to T-state (low O2 binding) from R-state (high O2 binding) because protonated His is bonded tightly to nearby Asp and Lys
• iron contained on the inside of heme, preventing free iron from reacting with O2 and allows binding of O2 in a controlled manner

  • histidine residue is a 5th point of coordination for iron
• structure of myoglobin is all helical (no beta sheets)
• hemoglobin is a tetromere

  • (αβ)2 -> 2 alpha and 2 beta chains
  • not entirely symmetrical
  • α2/β2 and α1/β1 have more intimate interaction than α2/β1 and α1/β2
  • α1 and α2, and β1 and β2 are diagonal
• binding properties of proteins

  • a ligand is defined as a molecule that binds reversibly to a protein. can be a small molecule (like O2) or a large molecule (another protein)
  • a ligand binds at a site on the protein called a binding site, which is complementary to the ligand in size, shape, charge, and hydrophobic or hydrophilic character. interaction is usually quite specific and selective
• ligand binding is transient; this allows the organism to respond rapidly and reversibly to changing metabolic and/or environmental circumstances

  • non covalent interactions usually reversible
• the equilibrium dissociation constant (Kd) is defined as the concentration of ligand (units=molar) at which 1/2 of the available ligand binding sites (on average) are occupied. the more tightly a protein binds a ligand, the lower the value of Kd

  • concentration in which 50% of binding sites occupied
• oxygen-binding curves for myoglobin and hemoglobin

  • curves shifted to the right signifies weaker binding
  • curves shifted to the left signifies tighter binding
  • myoglobin curves generally farther left than hemoglobin
  • carbon monoxide has a higher binding affinity to oxygen and shifts the curve to the left
• pH drops: protonate His, more interactions/weaker oxygen binding, curve shifts right
• the Bohr effect


  a decrease in pH of 0.8 units (7.6 to 6.8) doubles the ability of Hb to unload its O2 to Mb
• carbon monoxide poisoning

  • CO binds to Hb ˜ 250 times more tightly than does O2
  • a single bound CO stabilizes the R state of Hb, which increases affinity for O2 at the other three binding sites and prevents O2 from being delivered to the tissues (Hb can't go to T state)
  • earliest symptoms of CO poisoning are non-specific and readily confused with flu-like syndromes (headache, nausea, vertigo)
  • high concentrations of HbCO (50-60%) lead to seizures, coma and death
• Oxidation of Fe2+ and Fe3+ and formation of methemoglobin

  • exposure to exogenous oxidizing drugs (eg. benzocaine, dapsone) can increase rate of Fe3+ oxidation ˜ 1000 fold
  • Fe2+ -> Fe3+ inactivates hemoglobin (won't bind O2)
  • ingestion of compounds containing nitrates (bismuth nitrate) or well water contaminated by nitrates can also cause methemoglobinemia
  • there is an enzyme in RBCs that reduces Fe3+ back to Fe2+
• 2,3-biphosphoglycerate (BPG) is a side product of the glycolytic pathway

  • shifts binding curve to the right, making oxygen/Hb binding weaker
  • BPG concentration goes down with altitude