LEC 4 - ANTIBODIES / IMMUNOGLOBULINS

What are antibodies (immunoglobulins)?

They are specialized globular proteins that circulate in the blood and bind to harmful invaders called antigens

Why are antibodies also called immunoglobulins?

Because they are specialized globular proteins that play a vital role in immunity

What are antibodies made of?

They are glycoproteins composed of 86-98% polypeptide and 2-14% carbohydrate

What is the main humoral element of the adaptive immune response?

Antibodies (immunoglobulins)

What are the main functions of antibodies?

Antigen Recognition - Direct binding between antigen and antibody

Opsonization - Enhances or hastens phagocytosis

Complement Activation - Activates the complement system

Where are antibodies found?

On the surface of B cells and circulating in the blood

What is the role of antibodies on the surface of B cells?

They act as receptors that precisely identify foreign antigens to start an immune response

How are antibodies produced?

Antigen-induced stimulation of B lymphocytes leads to differentiation into plasma cells, which produce antibodies

What are gamma globulins?

They are immunoglobulins (antibodies) that appear in the γ region or γ band during protein electrophoresis

Why do immunoglobulins appear in the γ region?

Because they are the slowest-moving proteins and have a charge close to neutral

What is the basic structure of all immunoglobulin molecules?

A four-chain (tetrapeptide) unit made up of two heavy (H) chains and two light (L) chains

What are the two types of chains found in immunoglobulins?

Heavy (H) chains and Light (L) chains

How are the heavy and light chains held together?

By disulfide bonds

What is the heavy chain in an immunoglobulin molecule?

It is the larger chain of the molecule that extends the full length of the antibody

What is the light chain in an immunoglobulin molecule?

It is the smaller chain found on all immunoglobulin types but does not extend the full length of the molecule

What are the two main subtypes of light chains?

Kappa (κ) chains and Lambda (λ) chains

Which light chain subtype is more common?

Kappa (κ) chains, making up about 60% of light chains

What percentage of light chains are lambda (λ)?

About 40%

What are the two main regions of an antibody molecule?

The Variable Region and the Constant Region

Where is the variable region located?

At the amino terminal end of the antibody molecule

What is the function of the variable region?

It defines the specificity of an antibody for its antigen

How many amino acids make up the variable region?

The first 110 amino acids

Where is the constant region found?

At the carboxy terminal end of the antibody

From which amino acid does the constant region start?

It starts from the 111th amino acid onward for both heavy and light chains

Who first described the structure of the antibody?

Gerald Edelman and Rodney Porter

Which antibody type did Edelman and Porter use in their research?

The IgG antibody, the most prevalent in the body

What did Gerald Edelman use to separate immunoglobulins?

The analytic ultracentrifuge to separate based on molecular weight

What was the sedimentation coefficient of an intact IgG molecule?

7S

What two fractions did Edelman find when separating IgG?

3.5S - Heavy (H) chain (larger molecule)

2.2S - Light (L) chain (smaller portion)

What did Edelman conclude about the structure of immunoglobulins?

That each molecule has 2 heavy chains and 2 light chains (H₂L₂) — the generalized formula for all immunoglobulins

What enzyme did Rodney Porter use in his experiment?

The proteolytic enzyme papain

What did papain cleavage of IgG produce?

One Fc fragment (crystallizable fragment)

Two Fab fragments (antigen-binding fragments)

Where does papain cleave the IgG molecule?

Below the disulfide bonds that hold the two heavy chains together

What enzyme did Alfred Nisonoff use in his experiment?

Pepsin

How does pepsin cleave the IgG molecule?

At the carboxy terminal side of the interchain disulfide bonds

What fragments are produced by pepsin cleavage?

F(ab′)₂ fragment - retains all antigen-binding ability

Fc′ (Fc prime) fragment

What are the two main types of antibody fragments?

Fc fragment and Fab fragment

What does "Fc" stand for?

Fragment Crystallizable

Why is the Fc fragment called "crystallizable"?

Because it spontaneously crystallizes at 4°C

What part of the antibody does the Fc fragment represent?

The carboxy-terminal (tail) halves of the two heavy chains, consisting entirely of constant regions

What is the main function of the Fab fragment?

Antigen binding

What chains make up a Fab fragment?

One light chain and half of a heavy chain, held together by disulfide bonds

How many Fab fragments does an antibody have?

Two identical Fab fragments, each with one antigen-binding site

Where is the hinge region located in an antibody molecule?

Between the CH1 and CH2 regions of the heavy chain

What amino acid characteristics define the hinge region?

It is rich in hydrophobic residues and has a high proline content

What is the function of the hinge region?

It confers flexibility to the antibody molecule

Why is the hinge region susceptible to proteolysis?

Because of the presence of disulfide bonds

Which immunoglobulin has the longest hinge region?

IgD

Which immunoglobulin classes do not have a hinge region?

IgM and IgE

Where is the carbohydrate portion of an immunoglobulin attached?

In the CH2 domain of the heavy chain

What is one function of the carbohydrate portion in immunoglobulins?

It increases the solubility of the immunoglobulin molecule

How does the carbohydrate portion protect the antibody?

It provides protection against degradation

How does the carbohydrate portion affect the Fc domain?

It enhances the functional activity of the Fc domains

Does the basic four-chain structure of an antibody exist as a straight Y-shape?

No, it is folded into compact globular subunits

What is the three-dimensional structure of an immunoglobulin composed of?

Beta-pleated sheets (β-pleated structure)

What are hypervariable regions (HVRs) also known as?

Complementarity Determining Regions (CDRs)

Where are hypervariable regions located?

Within the variable regions of both heavy (H) and light (L) chains

What is the function of hypervariable regions (CDRs)?

They are responsible for the diversity of immunoglobulin molecules

What is the major immunoglobulin found in normal human serum?

IgG, comprising about 70-75% of total serum immunoglobulins

What is the sedimentation coefficient of IgG?

7S

Which immunoglobulin can cross the placenta?

IgG — the only one that can cross the placenta due to its small size

What does IgG's high diffusion coefficient indicate?

It can diffuse at a faster rate throughout tissues and fluids

When does IgG appear during an immune response?

t appears late in the immune response but persists longer because of its 23-day half-life

Which IgG subclass has the longest hinge region?

IgG3

Which IgG subclasses can cross the placenta?

All except IgG2

Which IgG subclass is most efficient in complement activation?

IgG3, followed by IgG1, IgG2, and IgG4 (3 → 1 → 2 → 4)

IgG3, followed by IgG1, IgG2, and IgG4 (3 → 1 → 2 → 4).

Passive natural immunity, as maternal IgG crosses the placenta to protect the infant

Is IgG better at precipitation or agglutination?

Precipitation, because it involves small soluble particles that are more easily brought together by the small IgG molecule

What is another name for IgM?

Macroglobulin

What is the sedimentation rate of IgM?

19S

How abundant is IgM in normal serum?

It is the 3rd most abundant immunoglobulin, comprising 5-10% of the total immunoglobulin pool

What type of antibody is IgM considered?

The primary response antibody

When does IgM appear and disappear in an immune response?

It is the first to appear and the first to disappear after antigenic stimulation, with a half-life of 6 days

When is IgM usually detected in infections?

During acute or current infections

Which antibody indicates past or recurring infections?

IgG

What are the two forms of IgM?

Pentameric form and Monomeric form

Where is the pentameric form of IgM found?

In the serum, secreted by plasma cells

How is the pentameric form of IgM structured?

It consists of five IgM monomers held together by a J (joining) chain

How many antigen-binding sites does the pentameric IgM have?

Ten (10) antigen-binding sites (Fab fragments)

What additional domain is present in IgM heavy chains?

An extra CH4 (Constant Heavy 4) domain

What is the monomeric form of IgM?

A single antibody molecule with a longer heavy chain, expressed as a membrane-bound antibody on B-cell surfaces

Why is IgM called the "agglutinating immunoglobulin"?

Because its pentameric structure provides 10 free antigen-binding sites, giving it high avidity for clumping antigens together

What is the predominant immunoglobulin in secretions?

IgA

What is the main function of IgA?

To protect mucosal surfaces from pathogens

Where is IgA commonly found?

In tears, saliva, milk, colostrum, and intestinal fluids

What percentage of the total immunoglobulin pool does IgA make up?

About 10-15%

What is the sedimentation coefficient of IgA?

7S

What are the two forms of IgA?

IgA1 (Monomeric IgA) and IgA2 (Dimeric or Secretory IgA)

Where is IgA1 (monomeric IgA) mainly found?

In the serum

What is the function of IgA1?

It helps downregulate phagocytosis, chemotaxis, bactericidal activity, and cytokine release

Where is IgA2 (dimeric/secretory IgA) mainly found?

In mucosal secretions such as saliva, tears, and intestinal fluids

What is the role of IgA2 at mucosal surfaces?

It prevents antigens from penetrating deeper into the body

What special component does IgA2 contain?

A secretory component (SC)

What is the function of the secretory component?

It makes IgA more resistant to proteolytic digestion and bacterial proteinases that can cleave IgA1

When and how was IgD first discovered?

In 1965, in a patient with multiple myeloma

How much of the total immunoglobulin pool does IgD make up?

Less than 0.001% of total immunoglobulins

What is the half-life of IgD?

About 1 to 3 days

Why is IgD extremely susceptible to proteolysis?

Because it has a very long hinge region containing 58 amino acids

Which antibody appears second during antigenic stimulation?

IgD, following IgM

Does secreted IgD serve a protective function in serum?

No, because it does not bind to complement, neutrophils, or macrophages, and does not cross the placenta