8536d_ch03_057-075 8/6/02 10:28 AM Page 62 mac79 Mac 79: 45_Bw Glasby et al. Immunology 5e 62 PART I Generation of B-Cell and T-Cell Response TABLE 3-3 Postulated mode of action of some commonly used adjuvants POSTULATED MODE OF ACTION Enhances Stimulates costimulator granuloma lymphocytes persistence formation nonspecifically Freunds incomplete adjuvant Freund's complete adjuvant +++ Aluminum potassium sulfate(alum) Mycobacterium tuberculosis ???+? +一 Bordetella pertussis Bacterial lipopolysaccharide( LPS) +++ Synthetic polynucleotides(poly IC/poly AU) Freunds complete adjuvant far more potent than the in- terminal portion, whereas the T cells responded only to epi- complete form. Activated macrophages are more phago- topes in the carboxyl-terminal portion cytic than unactivated macrophages and express higher Lymphocytes may interact with a complex antigen on sev- levels of class II MHC molecules and the membrane mole- eral levels of antigen structure. An epitope on a protein anti ules of the B7 family. The increased expression of class iI gen may involve elements of the primary, secondary, tertiary, MHC increases the ability of the antigen-presenting cell to and even quaternary structure of the protein(see Figure 3-1) present antigen to TH cells. B7 molecules on the antigen- In polysaccharides, branched chains are commonly present, presenting cell bind to CD28, a cell-surface protein on TH and multiple branches may contribute to the conformation cells, triggering co-stimulation, an enhancement of the T- of epitopes cell immune response. Thus, antigen presentation and the The recognition of antigens by t cells and B cells is funda requisite co-stimulatory signal usually are increased in the mentally different(Table 3-4). B cells recognize soluble anti- presence of adj gen when it binds to their membrane-bound antibody. Alum and Freund's adjuvants also stimulate a local, Because B cells bind antigen that is free in solution, the epi- chronic inflammatory response that attracts both phagocytes topes they recognize tend to be highly accessible sites on the and lymphocytes. This infiltration of cells at the site of the exposed surface of the immunogen. As noted previousl adjuvant injection often results in formation of a dense, most T cells recognize only peptides combined with MHC macrophage-rich mass of cells called a granuloma. Because molecules on the surface of antigen-presenting cells and al the macrophages in a granuloma are activated, this mecha- tered self-cells; T-cell epitopes, as a rule, cannot be consid nism also enhances the activation of TH cell ered apart from their associated MHC molecules Other adjuvants (e.g, synthetic polyribonucleotides and bacterial lipopolysaccharides)stimulate the nonspecific pro- Properties of B-Cell Epitopes Are Determined liferation of lymphocytes and thus increase the likelihood of by the Nature of the Antigen-Binding Site antigen-induced clonal selection of lymphocytes Several generalizations have emerged from studies in which the molecular features of the epitope recognized by B cells Epitopes have been established The ability to function as a B-cell epitope is determined by As mentioned in Chapter 1, immune cells do not interact the nature of the antigen-binding site on the antibody molecul with, or recognize, an entire immunogen molecule; instead, displayed by B cells. Antibody binds to an epitope by weak lymphocytes recognize discrete sites on the macromolecule noncovalent interactions operate only over called epitopes, or antigenic determinants. Epitopes are the tances. For a strong bond, the antibodys binding site and the mmunologically active regions of an immunogen that bind epitope must have complementary shapes that place the in- to secreted antibodies. Studies with small antigens have or teracting groups near each other. This requirement poses to antigen-specific membrane receptors on lymphocytes some restriction on the properties of the epitope. The size of realed that B and T cells recognize different epitopes on the the epitope recognized by a B cell can be no larger than the same antigenic molecule. For example, when mice were im- size of the antibody s binding site. For any given antigen-an munized with glucagon, a small human hormone of 29 tibody reaction, the shape of the epitope that can be recog amino acids, antibody was elicited to epitopes in the amino- nized by the antibody is determined by the shape assumed by
Freund’s complete adjuvant far more potent than the incomplete form. Activated macrophages are more phagocytic than unactivated macrophages and express higher levels of class II MHC molecules and the membrane molecules of the B7 family. The increased expression of class II MHC increases the ability of the antigen-presenting cell to present antigen to TH cells. B7 molecules on the antigenpresenting cell bind to CD28, a cell-surface protein on TH cells, triggering co-stimulation, an enhancement of the Tcell immune response. Thus, antigen presentation and the requisite co-stimulatory signal usually are increased in the presence of adjuvant. Alum and Freund’s adjuvants also stimulate a local, chronic inflammatory response that attracts both phagocytes and lymphocytes. This infiltration of cells at the site of the adjuvant injection often results in formation of a dense, macrophage-rich mass of cells called a granuloma. Because the macrophages in a granuloma are activated, this mechanism also enhances the activation of TH cells. Other adjuvants (e.g., synthetic polyribonucleotides and bacterial lipopolysaccharides) stimulate the nonspecific proliferation of lymphocytes and thus increase the likelihood of antigen-induced clonal selection of lymphocytes. Epitopes As mentioned in Chapter 1, immune cells do not interact with, or recognize, an entire immunogen molecule; instead, lymphocytes recognize discrete sites on the macromolecule called epitopes, or antigenic determinants. Epitopes are the immunologically active regions of an immunogen that bind to antigen-specific membrane receptors on lymphocytes or to secreted antibodies. Studies with small antigens have revealed that B and T cells recognize different epitopes on the same antigenic molecule. For example, when mice were immunized with glucagon, a small human hormone of 29 amino acids, antibody was elicited to epitopes in the aminoterminal portion, whereas the T cells responded only to epitopes in the carboxyl-terminal portion. Lymphocytes may interact with a complex antigen on several levels of antigen structure. An epitope on a protein antigen may involve elements of the primary, secondary, tertiary, and even quaternary structure of the protein (see Figure 3-1). In polysaccharides, branched chains are commonly present, and multiple branches may contribute to the conformation of epitopes. The recognition of antigens by T cells and B cells is fundamentally different (Table 3-4). B cells recognize soluble antigen when it binds to their membrane-bound antibody. Because B cells bind antigen that is free in solution, the epitopes they recognize tend to be highly accessible sites on the exposed surface of the immunogen. As noted previously, most T cells recognize only peptides combined with MHC molecules on the surface of antigen-presenting cells and altered self-cells; T-cell epitopes, as a rule, cannot be considered apart from their associated MHC molecules. Properties of B-Cell Epitopes Are Determined by the Nature of the Antigen-Binding Site Several generalizations have emerged from studies in which the molecular features of the epitope recognized by B cells have been established. The ability to function as a B-cell epitope is determined by the nature of the antigen-binding site on the antibody molecules displayed by B cells. Antibody binds to an epitope by weak noncovalent interactions, which operate only over short distances. For a strong bond, the antibody’s binding site and the epitope must have complementary shapes that place the interacting groups near each other. This requirement poses some restriction on the properties of the epitope. The size of the epitope recognized by a B cell can be no larger than the size of the antibody’s binding site. For any given antigen-antibody reaction, the shape of the epitope that can be recognized by the antibody is determined by the shape assumed by 62 PART II Generation of B-Cell and T-Cell Responses TABLE 3-3 Postulated mode of action of some commonly used adjuvants POSTULATED MODE OF ACTION Prolongs Enhances Induces Stimulates antigen co-stimulatory granuloma lymphocytes Adjuvant persistence signal formation nonspecifically Freund’s incomplete adjuvant Freund’s complete adjuvant Aluminum potassium sulfate (alum) ? Mycobacterium tuberculosis ? Bordetella pertussis ? Bacterial lipopolysaccharide (LPS) Synthetic polynucleotides (poly IC/poly AU) ? 8536d_ch03_057-075 8/6/02 10:28 AM Page 62 mac79 Mac 79:45_BW:Goldsby et al. / Immunology 5e:
8536ach03057-0758/7/029:18 AM Page63mac79Mac79:45Bw: Goosby et Immunology 5e: Antigens CHAPTER 3 TABLE 3-4 Comparison of antigen recognition by T cells and B cells Characteristic B cells T cells Interaction with antigen Involves binary complex of membrane Involves ternary complex of T-cell receptor, Ag, lg and Ag and MHC molecule Binding of soluble antigen Involvement of mhc molecules Required to display processed antigen de, lipid les Accessible, hydrophilic, mobile peptide Internal linear peptides produced by containing sequential or nonsequential processing of antigen and bound to amino acids MHC molecules the sequences of amino acids in the binding site and the that make up the binding site. Despite differences in the chemical environment that they produce. binding patterns of small haptens and large antigens, Chap Smaller ligands such as carbohydrates, small oligonu- ter 4 will show that all antibody binding sites are assembled cleotides, peptides, and haptens often bind within a deep from the same regions of the antibody molecule--namely, pocket of an antibody. For example, angiotensin Il, a small parts of the variable regions of its polypeptide chains octapeptide hormone, binds within a deep and narrow horne(725 A2)of a monoclonal antibody specific for the hormone(Figure 3-2). Within this groove, the bound pep- tide hormone folds into a compact structure with two turns, which brings its amino(N-terminal) and carboxyl(C-termi nal)termini close together. All eight amino acid residues of the octapeptide are in van der Waals contact with 14 residues of the antibodys groove. a quite different picture of epitope structure emerges from x-ray crystallographic analyses of monoclonal antibod ies bound to globular protein antigens such as hen egg-white lysozyme(HEL)and neuraminidase(an envelope glycopro tein of influenza virus). These antibodies make contact with the antigen across a large flat face(Figure 3-3). The interact- ing face between antibody and epitope is a flat or undulating surface in which protrusions on the epitope or antibody are matched by corresponding depressions on the antibody or epitope. These studies have revealed that 15-22 amino acids on the surface of the antigen make contact with a similar number of residues in the antibodys binding site; the surface area of this large complementary interface is between 650 A and 900 A2. For these globular protein antigens, then,the shape of the epitope is entirely determined by the tertiary conformation of the native protein. Thus, globular protein antigens and small peptide anti- gens interact with antibody in different ways( Figure 3-4). Typically, larger areas of protein antigens are engaged by the antibody binding site. In contrast, a small peptide such as an- giotensin II can fold into a compact structure that occupies less space and fits into a pocket or cleft of the binding site. This pattern is not unique to small peptides; it extends to the FIGURE 3.2 Three-dimensional structure of an octapeptide hor binding of low-molecular-weight antigens of various chemi- mone(angiotensin Il)complexed with a monoclonal antibody Fab I types. However, these differences between the binding of fragment, the antigen-binding unit of the antibody molecule. The an- small and large antigenic determinants do not reflect funda- giotensin ll peptide is shown in red, the heavy chain in blue, and the mental differences in the regions of the antibody molecule light chain in purple. From K C Garcia et al., 1992, Science 257: 502.1
the sequences of amino acids in the binding site and the chemical environment that they produce. Smaller ligands such as carbohydrates, small oligonucleotides, peptides, and haptens often bind within a deep pocket of an antibody. For example, angiotensin II, a small octapeptide hormone, binds within a deep and narrow groove (725 Å2 ) of a monoclonal antibody specific for the hormone (Figure 3-2). Within this groove, the bound peptide hormone folds into a compact structure with two turns, which brings its amino (N-terminal) and carboxyl (C-terminal) termini close together. All eight amino acid residues of the octapeptide are in van der Waals contact with 14 residues of the antibody’s groove. A quite different picture of epitope structure emerges from x-ray crystallographic analyses of monoclonal antibodies bound to globular protein antigens such as hen egg-white lysozyme (HEL) and neuraminidase (an envelope glycoprotein of influenza virus). These antibodies make contact with the antigen across a large flat face (Figure 3-3). The interacting face between antibody and epitope is a flat or undulating surface in which protrusions on the epitope or antibody are matched by corresponding depressions on the antibody or epitope. These studies have revealed that 15–22 amino acids on the surface of the antigen make contact with a similar number of residues in the antibody’s binding site; the surface area of this large complementary interface is between 650 Å2 and 900 Å2 . For these globular protein antigens, then, the shape of the epitope is entirely determined by the tertiary conformation of the native protein. Thus, globular protein antigens and small peptide antigens interact with antibody in different ways (Figure 3-4). Typically, larger areas of protein antigens are engaged by the antibody binding site. In contrast, a small peptide such as angiotensin II can fold into a compact structure that occupies less space and fits into a pocket or cleft of the binding site. This pattern is not unique to small peptides; it extends to the binding of low-molecular-weight antigens of various chemical types. However, these differences between the binding of small and large antigenic determinants do not reflect fundamental differences in the regions of the antibody molecule that make up the binding site. Despite differences in the binding patterns of small haptens and large antigens, Chapter 4 will show that all antibody binding sites are assembled from the same regions of the antibody molecule—namely, parts of the variable regions of its polypeptide chains. Antigens CHAPTER 3 63 TABLE 3-4 Comparison of antigen recognition by T cells and B cells Characteristic B cells T cells Interaction with antigen Involves binary complex of membrane Involves ternary complex of T-cell receptor, Ag, Ig and Ag and MHC molecule Binding of soluble antigen Yes No Involvement of MHC molecules None required Required to display processed antigen Chemical nature of antigens Protein, polysaccharide, lipid Mostly proteins, but some lipids and glycolipids presented on MHC-like molecules Epitope properties Accessible, hydrophilic, mobile peptides Internal linear peptides produced by containing sequential or nonsequential processing of antigen and bound to amino acids MHC molecules FIGURE 3-2 Three-dimensional structure of an octapeptide hormone (angiotensin II) complexed with a monoclonal antibody Fab fragment, the antigen-binding unit of the antibody molecule. The angiotensin II peptide is shown in red, the heavy chain in blue, and the light chain in purple. [From K. C. Garcia et al., 1992, Science 257:502.] 8536d_ch03_057-075 8/7/02 9:18 AM Page 63 mac79 Mac 79:45_BW:Goldsby et al. / Immunology 5e: