HAEMOGLOBIN (Hb)

Structure of Hb - Champe and Harvey 2nd ed.

Hb is a globular protein occurring in high concentration in red blood cells. Their main
function is to bind oxygen in the lungs and transport it to tissues and cells, and to
transport carbon dioxide and protons from the tissues to the lungs.

Types:
HbA is the major adult Hb, consisting of four polypeptide chains (2 and 2) held
together by non-covalent interactions.
HbA2 is a minor adult Hb, consisting of 2 and 2 chains
HbF is the fetal form of Hb, consisting of 2 and 2 chains.

Heme:

Hemeproteins are specialised proteins that contain heme as the prosthetic group.
Heme is a porphyrin molecule (definition : able to combine with metallic elements
such as Mg, Cu and Fe) containing an iron (Fe) atom at its center.
This Fe is in the ferrous (2+) oxidation state in functional Hb.

Fe2+ can form six ligand bonds. Four bound to pyrrole nitrogen atoms of the
porphyrin, the fifth and sixth are directed along an axis perpendicular to the plane of
the porphyrin ring. The fifth bond is to the nitrogen of a histidine imidazole. The sixth
co-ordinate bond may be to oxygen (oxygenated state) or unoccupied (deoxy state).

Therefore Fe2+ or heme binds one oxygen atom.

Structure of heme - Champe and Harvey 2nd ed

Quarternary structure
Hb consists of 2 identical dimers - ()1 and ()2
Within each dimer polypeptide chains are held tightly together by hydrophobic
interactions, as well as ionic bonds and hydrogen bonds.
The two dimers are held together by polar bonds.
The two dimers are able to move relative to each other and therefore occupy different
positions in the oxygenated/deoxygenated state.

Schematic diagram showing the structural changes resulting from oxygenation and
deoxygenation of Hb - Champe and harvey 2nd ed.

Tense/taut form = deoxygenated state - dimers interact through ionic and hydrogen
bonds, which constrain movement of polypeptide chains.
Relaxed form = oxygenated state - binding of oxygen to one site ruptures some of
the ionic/ hydrogen bonds between the dimers. This increases the affinity for oxygen
and facilitates further binding of this atom.
Binding of oxygen
Hb can bind 4 molecules of oxygen, one at each heme group.
% saturation can vary between 0-100%.
A measure of saturation at different partial pressures of oxygen gives the oxygen
dissociation curve.
This curve is sigmoidal in shape, which indicates that co-operative binding of oxygen
occurs, i.e., binding of one oxygen molecule at one heme group increases affinity of
remaining heme groups for oxygen.
PLASMA PROTEINS

Plasma consists of water, electrolytes, metabolites, nutrients, proteins and

hormones.
The concentration of total protein is 7-7.5 g/dL. Proteins are a major source of solids
in the plasma.

Six generalisations about plasma proteins

1. Most are synthesised in the liver, but the gamma-globulins are synthesised in
plasma cells and others may be synthesised in endothelial cells.
2. Most are synthesised on membrane-bound polyribosomes and then secreted via
the following route in the plasma: rER, sER, Golgi, plasma mambrane. They are
synthesised as preproteins and initially contain N-terminal signal peptides, which
are subjected to post-translational modifications (proteolysis, glycosylation,
phosphorylation, etc).
3. Almost all are glycoproteins, containing either O- or N-linked oligosaccharide
chains. Albumin contains no sugar residues, and is therefore the exception.
4. May exhibit polymorphisms
5. Have a characteristic half-life in circulation.
6. Levels increase during acute inflammatory states or secondary to certain types of
tissue damage. They therefore play a role in the body's response to inflammation.

HUMAN PLASMA PROTEINS

Albumin
Albumin is the major protein in human plasma, making up 50-60% 0f the total plasma
protein.
Produced in the liver at the rate of 12g/day, representing 25% of hepatic protein
synthesis and half the total secreted protein.
Synthesised as a preprotein, the signal peptide is removed as it passes into rER.
Mature albumin consists of one polypeptide chain of 585 amino acids and 17
disulphide bonds.
MW = 69 000, 3 domains
Ellipsoidal shape, i.e., does not increase the viscosity of plasma as much as the
elongated molecule.
Functions: - control of osmotic pressure of plasma
- binding of various ligands, including freefatty acids, calcium, bilirubin,
steroid hormones etc.
- transport of copper
- transport of drugs, eg. Penicillin, aspirin

Haptoglobin
Haptoglobin (Hp) is a plasma protein that binds extra0corpuscular Hb in a tight non-
covalent complex.
40-180 mg Hb-bibding ability per dL.
MW Hp = 90 000, MW Hb = 65 000, total MW Hp-Hb = 155 000
Free Hb passes through the glomerulus into the tubules and precipitates. The Hp-Hb
complex is too large to traverse the glomerulus. Hp therefore prevents loss of free Hb
in the kidney.
Low levels in patients with hemolytic anemia.
3 polymorphic forms: Hp 1-1, Hp 2-1, Hp 2-2

Transferrin
Transferrin is 1-globulin with a molecular weight of 80 000
Is a glycoprotein that is synthesised in the liver.
20 polymorphic forms exist
concentration = 300 mg/dL which binds 300 g Fe/dL = total Fe-binding capacity of
plasma.
Function: - Transports Fe in circulation to the site of need, thereby decreasing the
potential toxicity of Fe.
Mechanism of action: Transferrin binds to its cell receptors, is internalised by
receptor-mediated endocytosis and enters the lysosome. The acid pH causes
dissociation of Fe, transferrin is returned to the plasma membrane, dissociates from
its receptor and re-enters the plasma to pick up more Fe.

Ceruloplasmin
2- globulin with MW = 160 000
concentration = 30 mg/dL
synthesised in the liver, therefore concentration decreases in liver disease
Function: - transport copper
Can bind 6 atoms of copper, but this is not easily exchangable
About 10% of Cu is transported by albumin.

1-Antitrypsin
It is a single chain protein with 394 amino acids and 3 oligosaccharide chains.
Synthesised by hepatocytes and macrophages
MW = 52 000
Polymorphic
Function: - Inhibits serine proteases and others, also elastase and trypsin]
Plays a major clinical role in emphysema, where concentration is decreased due to a
polymorphism (PiZZ) and smoking (methionine residue 358 is oxidised to methionine
sulphoxide, which inactivates 1-antitrypsin.

Immunoglobulins

The immune system consists of 2 components, B and T lymphocytes. T-cells are

invloved in cell-mediated immunity, while B-cells synthesise humoral antibodies or
immunoglobulins (Ig).

All Ig consist of 2 identical light and 2 identical heavy chains held together as a
tetramer by disulphide bonds. Each chain is further subdivided into domains with
functional and structural significance
- light chain -- half at carboxyl terminal = constant region (CL)
-- half at the amino terminal = variable region (VL)
- heavy chain -- quarter at amino terminal = variable region (VH)
-- three quarters at carboxyl terminal = constant region (CH)
The portion that binds specific antigens is formed by the amino terminal portions or
variable regions.

2 types of light chains viz. Kappa and lambda, which are distinguished based on their
CL regions
five classes of h-chain have been found in humans, distinguished based on
differences in their CH regions.

The type of H-chain determines the class of Ig and thus its effector function.
Five classes of Ig exist viz., IgG, IgA, IgM, IgD, and IgE.

Lipoproteins
Lipoproteins are complexes of lipids and apolipoproteins.
They include chylomicrons, very low density lipoproteins (VLDL), low density
lipoproteins (LDL) and high density lipoproteins (HDL).
Chylomicrons have the lowest density and the largest size (more lipid, less protein)
VLDL and LDL are successively more dense, having less lipid and more protein.
HDL is the most dense.
Composition: neutral lipid core (triacylglycerol or cholesterol esters)
Function: - to keep lipids soluble as they transport them in the plasma (are
oriented so that their polar portions are exposed, rendering the particle
soluble in aqueous solution).
- to provide a mechanism of delivery of lipids to tissue.