NCBI Bookshelf. A service of the National Library of Medicine, National Institutes of Health.

Janeway CA Jr, Travers P, Walport M, et al. Immunobiology: The Immune System in Health and Disease. 5th edition. New York: Garland Science; 2001.

  • By agreement with the publisher, this book is accessible by the search feature, but cannot be browsed.
Cover of Immunobiology

Immunobiology: The Immune System in Health and Disease. 5th edition.

Show details

The structure of a typical antibody molecule

Antibodies are the secreted form of the B-cell receptor. An antibody is identical to the B-cell receptor of the cell that secretes it except for a small portion of the C-terminus of the heavy-chain constant region. In the case of the B-cell receptor the C-terminus is a hydrophobic membrane-anchoring sequence, and in the case of antibody it is a hydrophilic sequence that allows secretion. Since they are soluble, and secreted in large quantities, antibodies are easily obtainable and easily studied. For this reason, most of what we know about the B-cell receptor comes from the study of antibodies.

Antibody molecules are roughly Y-shaped molecules consisting of three equal-sized portions, loosely connected by a flexible tether. Three schematic representations of antibody structure, which has been determined by X-ray crystallography, are shown in Fig. 3.1. The aim of this part of the chapter is to explain how this structure is formed and how it allows antibody molecules to carry out their dual tasks—binding on the one hand to a wide variety of antigens, and on the other hand to a limited number of effector molecules and cells. As we will see, each of these tasks is carried out by separable parts of the molecule. The two arms of the Y end in regions that vary between different antibody molecules, the V regions. These are involved in antigen binding, whereas the stem of the Y, or the C region, is far less variable and is the part that interacts with effector cells and molecules.

Figure 3.1. Structure of an antibody molecule.

Figure 3.1

Structure of an antibody molecule. Panel a illustrates a ribbon diagram based on the X-ray crystallographic structure of an IgG antibody, showing the course of the backbones of the polypeptide chains. Three globular regions form a Y. The two antigen-binding (more...)

All antibodies are constructed in the same way from paired heavy and light polypeptide chains, and the generic term immunoglobulin is used for all such proteins. Within this general category, however, five different classes of immunoglobulinsIgM, IgD, IgG, IgA, and IgE—can be distinguished by their C regions, which will be described more fully in Chapter 4. More subtle differences confined to the V region account for the specificity of antigen binding. We will use the IgG antibody molecule as an example to describe the general structural features of immunoglobulins.

3-1. IgG antibodies consist of four polypeptide chains

IgG antibodies are large molecules, having a molecular weight of approximately 150 kDa, composed of two different kinds of polypeptide chain. One, of approximately 50 kDa, is termed the heavy or H chain, and the other, of 25 kDa, is termed the light or L chain (Fig. 3.2). Each IgG molecule consists of two heavy chains and two light chains. The two heavy chains are linked to each other by disulfide bonds and each heavy chain is linked to a light chain by a disulfide bond. In any given immunoglobulin molecule, the two heavy chains and the two light chains are identical, giving an antibody molecule two identical antigen-binding sites (see Fig. 3.1), and thus the ability to bind simultaneously to two identical structures.

Figure 3.2. Immunoglobulin molecules are composed of two types of protein chain: heavy chains and light chains.

Figure 3.2

Immunoglobulin molecules are composed of two types of protein chain: heavy chains and light chains. Each immunoglobulin molecule is made up of two heavy chains (green) and two light chains (yellow) joined by disulfide bonds so that each heavy chain is (more...)

Two types of light chain, termed lambda (λ) and kappa (κ), are found in antibodies. A given immunoglobulin either has κ chains or λ chains, never one of each. No functional difference has been found between antibodies having λ or κ light chains, and either type of light chain may be found in antibodies of any of the five major classes. The ratio of the two types of light chain varies from species to species. In mice, the average κ to λ ratio is 20:1, whereas in humans it is 2:1 and in cattle it is 1:20. The reason for this variation is unknown. Distortions of this ratio can sometimes be used to detect the abnormal proliferation of a clone of B cells. These would all express the identical light chain, and thus an excess of λ light chains in a person might indicate the presence of a B-cell tumor producing λ chains.

By contrast, the class, and thus the effector function, of an antibody, is defined by the structure of its heavy chain. There are five main heavy-chain classes or isotypes, some of which have several subtypes, and these determine the functional activity of an antibody molecule. The five major classes of immunoglobulin are immunoglobulin M (IgM), immunoglobulin D (IgD), immunoglobulin G (IgG), immunoglobulin A (IgA), and immunoglobulin E (IgE). Their heavy chains are denoted by the corresponding lower-case Greek letter (μ, δ, γ, α, and ε, respectively). IgG is by far the most abundant immunoglobulin and has several subclasses (IgG1, 2, 3, and 4 in humans). Their distinctive functional properties are conferred by the carboxy-terminal part of the heavy chain, where it is not associated with the light chain. We will describe the structure and functions of the different heavy-chain isotypes in Chapter 4. The general structural features of all the isotypes are similar and we will consider IgG, the most abundant isotype in plasma, as a typical antibody molecule.

3-2. Immunoglobulin heavy and light chains are composed of constant and variable regions

The amino acid sequences of many immunoglobulin heavy and light chains have been determined and reveal two important features of antibody molecules. First, each chain consists of a series of similar, although not identical, sequences, each about 110 amino acids long. Each of these repeats corresponds to a discrete, compactly folded region of protein structure known as a protein domain. The light chain is made up of two such immunoglobulin domains, whereas the heavy chain of the IgG antibody contains four (see Fig. 3.1a). This suggests that the immunoglobulin chains have evolved by repeated duplication of an ancestral gene corresponding to a single domain.

The second important feature revealed by comparisons of amino acid sequences is that the amino-terminal sequences of both the heavy and light chains vary greatly between different antibodies. The variability in sequence is limited to approximately the first 110 amino acids, corresponding to the first domain, whereas the remaining domains are constant between immunoglobulin chains of the same isotype. The amino-terminal variable or V domains of the heavy and light chains (VH and VL, respectively) together make up the V region of the antibody and confer on it the ability to bind specific antigen, while the constant domains (C domains) of the heavy and light chains (CH and CL, respectively) make up the C region (see Fig. 3.1b, c). The multiple heavy-chain C domains are numbered from the amino-terminal end to the carboxy terminus, for example CH1, CH2, and so on.

3-3. The antibody molecule can readily be cleaved into functionally distinct fragments

The protein domains described above associate to form larger globular domains. Thus, when fully folded and assembled, an antibody molecule comprises three equal-sized globular portions joined by a flexible stretch of polypeptide chain known as the hinge region (see Fig. 3.1b). Each arm of this Y-shaped structure is formed by the association of a light chain with the amino-terminal half of a heavy chain, whereas the trunk of the Y is formed by the pairing of the carboxy-terminal halves of the two heavy chains. The association of the heavy and light chains is such that the VH and VL domains are paired, as are the CH1 and CL domains. The CH3 domains pair with each other but the CH2 domains do not interact; carbohydrate side chains attached to the CH2 domains lie between the two heavy chains. The two antigen-binding sites are formed by the paired VH and VL domains at the ends of the two arms of the Y (see Fig. 3.1b).

Proteolytic enzymes (proteases) that cleave polypeptide sequences have been used to dissect the structure of antibody molecules and to determine which parts of the molecule are responsible for its various functions. Limited digestion with the protease papain cleaves antibody molecules into three fragments (Fig. 3.3). Two fragments are identical and contain the antigen-binding activity. These are termed the Fab fragments, for Fragment antigen binding. The Fab fragments correspond to the two identical arms of the antibody molecule, which contain the complete light chains paired with the VH and CH1 domains of the heavy chains. The other fragment contains no antigen-binding activity but was originally observed to crystallize readily, and for this reason was named the Fc fragment, for Fragment crystallizable. This fragment corresponds to the paired CH2 and CH3 domains and is the part of the antibody molecule that interacts with effector molecules and cells. The functional differences between heavy-chain isotypes lie mainly in the Fc fragment.

Figure 3.3. The Y-shaped immunoglobulin molecule can be dissected by partial digestion with proteases.

Figure 3.3

The Y-shaped immunoglobulin molecule can be dissected by partial digestion with proteases. Papain cleaves the immunoglobulin molecule into three pieces, two Fab fragments and one Fc fragment (upper panels). The Fab fragment contains the V regions and (more...)

The protein fragments obtained after proteolysis are determined by where the protease cuts the antibody molecule in relation to the disulfide bonds that link the two heavy chains. These lie in the hinge region between the CH1 and CH2 domains and, as illustrated in Fig. 3.3, papain cleaves the antibody molecule on the amino-terminal side of the disulfide bonds. This releases the two arms of the antibody as separate Fab fragments, whereas in the Fc fragment the carboxy-terminal halves of the heavy chains remain linked.

Another protease, pepsin, cuts in the same general region of the antibody molecule as papain but on the carboxy-terminal side of the disulfide bonds (see Fig. 3.3). This produces a fragment, the F(ab′)2 fragment, in which the two antigen-binding arms of the antibody molecule remain linked. In this case the remaining part of the heavy chain is cut into several small fragments. The F(ab′)2 fragment has exactly the same antigen-binding characteristics as the original antibody but is unable to interact with any effector molecule. It is thus of potential value in therapeutic applications of antibodies as well as in research into the functional role of the Fc portion.

Genetic engineering techniques also now permit the construction of many different antibody-related molecules. One important type is a truncated Fab comprising only the V domain of a heavy chain linked by a stretch of synthetic peptide to a V domain of a light chain. This is called single-chain Fv, named from Fragment variable. Fv molecules may become valuable therapeutic agents because of their small size, which allows them to penetrate tissues readily. They can be coupled to protein toxins to yield immunotoxins with potential application, for example, in tumor therapy in the case of a Fv specific for a tumor antigen (see Chapter 14).

3-4. The immunoglobulin molecule is flexible, especially at the hinge region

The hinge region that links the Fc and Fab portions of the antibody molecule is in reality a flexible tether, allowing independent movement of the two Fab arms, rather than a rigid hinge. This has been demonstrated by electron microscopy of antibodies bound to haptens. These are small molecules of various sorts, typically about the size of a tyrosine side chain. They can be recognized by antibody but are only able to stimulate production of antihapten antibodies when linked to a larger protein carrier(see Appendix I, Section A-1). An antigen made of two identical hapten molecules joined by a short flexible region can link two or more anti-hapten antibodies, forming dimers, trimers, tetramers, and so on, which can be seen by electron microscopy (Fig. 3.4). The shapes formed by these complexes demonstrate that antibody molecules are flexible at the hinge region. Some flexibility is also found at the junction between the V and C domains, allowing bending and rotation of the V domain relative to the C domain. For example, in the antibody molecule shown in Fig. 3.1a, not only are the two hinge regions clearly bent differently, but the angle between the V and C domains in each of the two Fab arms is also different. This range of motion has led to the junction between the V and C domains being referred to as a ‘molecular balland-socket joint.’ Flexibility at both the hinge and V-C junction enables the binding of both arms of an antibody molecule to sites that are various distances apart, for example, sites on bacterial cell-wall polysaccharides. Flexibility at the hinge also enables the antibodies to interact with the antibody-binding proteins that mediate immune effector mechanisms.

Figure 3.4. Antibody arms are joined by a flexible hinge.

Figure 3.4

Antibody arms are joined by a flexible hinge. An antigen consisting of two hapten molecules (red balls in diagrams) that can cross-link two antigen-binding sites is used to create antigen:antibody complexes, which can be seen in the electron micrograph. (more...)

3-5. The domains of an immunoglobulin molecule have similar structures

As we saw in Section 3-2, immunoglobulin heavy and light chains are composed of a series of discrete protein domains. These protein domains all have a similar folded structure. Within this basic three-dimensional structure, there are distinct differences between V and C domains. The structural similarities and differences can be seen in the diagram of a light chain in Fig. 3.5. Each domain is constructed from two β sheets, which are elements of protein structure made up of strands of the polypeptide chain (β strands) packed together; the sheets are linked by a disulfide bridge and together form a roughly barrel-shaped structure, known as a β barrel. The distinctive folded structure of the immunoglobulin protein domain is known as the immunoglobulin fold.

Figure 3.5. The structure of immuno-globulin constant and variable domains.

Figure 3.5

The structure of immuno-globulin constant and variable domains. The upper panels show schematically the folding pattern of the constant (C) and variable (V) domains of an immunoglobulin light chain. Each domain is a barrel-shaped structure in which strands (more...)

Both the essential similarity of V and C domains and the critical difference between them are most clearly seen in the bottom panels of Fig. 3.5, where the cylindrical domains are opened out to reveal how the polypeptide chain folds to create each of the β sheets and how it forms flexible loops as it changes direction. The main difference between the V and C domains is that the V domain is larger, with an extra loop. We will see in Section 3-6 that the flexible loops of the V domains form the antigen-binding site of the immunoglobulin molecule.

Many of the amino acids that are common to C and V domains of immuno-globulin chains lie in the core of the immunoglobulin fold and are critical to its stability. For that reason, other proteins having sequences similar to those of immunoglobulins are believed to form domains of similar structure, and in many cases this has been demonstrated by crystallography. These immunoglobulin-like domains are present in many other proteins of the immune system, and in proteins involved in cell-cell recognition in the nervous system and other tissues. Together with the immunoglobulins and the T-cell receptors, they make up the extensive immunoglobulin superfamily.

Summary

The IgG antibody molecule is made up of four polypeptide chains, comprising two identical light chains and two identical heavy chains, and can be thought of as forming a flexible Y-shaped structure. Each of the four chains has a variable (V) region at its amino terminus, which contributes to the antigen-binding site, and a constant (C) region, which determines the isotype. The isotype of the heavy chain determines the functional properties of the antibody. The light chains are bound to the heavy chains by many noncovalent interactions and by disulfide bonds, and the V regions of the heavy and light chains pair in each arm of the Y to generate two identical antigen-binding sites, which lie at the tips of the arms of the Y. The possession of two antigen-binding sites allows antibody molecules to cross-link antigens and to bind them much more stably. The trunk of the Y, or Fc fragment, is composed of the carboxy-terminal domains of the heavy chains. Joining the arms of the Y to the trunk are the flexible hinge regions. The Fc fragment and hinge regions differ in antibodies of different isotypes, thus determining their functional properties. However, the overall organization of the domains is similar in all isotypes.

By agreement with the publisher, this book is accessible by the search feature, but cannot be browsed.

Copyright © 2001, Garland Science.
Bookshelf ID: NBK27144

Views

  • Cite this Page
  • Disable Glossary Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...