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Immunology -Chapter 4: Antigen and Antibody Structure
Immunology -Chapter 4: Antigen and Antibody Structure. Antigens that cause a strong immune response can have very diverse chemical structures. Antibodies are antigen-specific proteins produced when B cells are in contact with antigens and circulate in the blood and lymph as plasma components. Each individual can synthesize a large number of different antibody molecules and interact with specific antigens.
The first part of this chapter introduces the various types of antigens and which antigens can cause the best immune response. The second part describes the general structure of various types of human antibodies (immunoglobulins) and some molecular biological characteristics of the antibodies. Chapter 5 explores the nature of antibody-antigen interactions in more depth, and Chapter 6 explains how antibody diversity is produced.
Antigens that cause a strong immune response can have very diverse chemical structures. Antibodies are antigen-specific proteins produced when B cells are in contact with antigens and circulate in the blood and lymph as plasma components. Each individual can synthesize a large number of different antibody molecules and interact with specific antigens.
2.1 Types of antigens
In the first chapter, antigens are referred to as substances recognized by the adaptive immune system receptors. Antigens can be classified according to the extent to which they induce an immune response and their source.
An immunogen is a substance that itself can cause an immune response, such as the production of antibodies. Effective immunogens are large in size, with a molecular weight generally greater than 6000, and have complex chemical structures. For example, a protein immunogen composed of 20 different amino acid residues is more immunogenic than a nucleic acid composed of 4 different nucleotide bases. The surface antigen of hepatitis B virus is the immunogen (Box 4.1).
In contrast, haptens are compounds that can be bound by immune receptors, but they do not necessarily trigger an immune response by themselves. For example, relatively simple compounds like penicillin cannot cause antibody responses by themselves. If the hapten is coupled to a large molecule such as a protein, an antibody that can bind to the hapten very specifically can be produced (Fig 4.1).
Fig 4.1丨Hapten-carrier conjugate
The third antigen is the tolerance antigen. These molecules are recognized by the adaptive immune system, but the adaptive immune system does not respond.
There are different sources of antigens. Antigens from pathogens are usually used as immunogens or haptens to cause a strong immune response. Proteins from others-such as the different major histocompatibility complex (MHC) alleles that appear in kidney transplants-can act as allogeneic antigens, triggering a strong immune response. Antigens derived from food (such as peanuts) can be used as tolerogens for most people. When tolerance procedures go wrong, these antigens may act as allergens and trigger harmful immune responses, and lack of tolerance may also cause normal host molecules to become self-antigens.
An epitope is a part of an antigen, such as a viral protein, to which antibodies can react. Viral proteins may contain a large number of epitopes that can interact with many different specific antibodies or T cell receptors. As shown in Fig 4.2, there are usually two types of epitopes: one is discontinuous epitopes or conformational epitopes. This epitope is the aggregation of amino acid residues in discontinuous regions of the polypeptide chain into a three-dimensional (3 -D) Shaped structure, the other is continuous epitopes, also called linear epitopes, which are continuous regions of the sequence, such as amino acids 12 to 22 in the polypeptide chain.
Recognition of linear epitopes by immune receptors on T cells requires processed antigens to be presented to them (related to MHC molecules, see Chapter 10). As shown in Fig 4.2, antibodies can recognize these two types of epitopes.
Fig 4.2 | Linear epitopes and discontinuous epitopes.
3.1 Separation and identification of antibodies
Antibodies are proteins that specifically react with the antigen that stimulates their production. The two key characteristics of antibodies are that they are highly specific and only bind to specific antigens, and they have a high affinity, which means that they bind very tightly to the antigen.
Antibodies account for about 20% of plasma proteins. They were initially detected by analytical techniques such as electrophoresis. In electrophoresis, individual blood proteins are separated according to their charge and molecular weight. During the electrophoresis process, the antibody will be separated into the gamma region. The antibody is sometimes called the gamma globulin in the serum (Fig 4.3).
Fig 4.3 | Human serum total electrophoresis Ig, immunoglobulin
Generally, the part of serum containing antibodies is called immunoglobulin. Since normal serum immunoglobulin is a heterogeneous product of many B cell clones (polyclonal immunoglobulin [Ig]), the immunoglobulin in serum constitutes a highly heterogeneous protein spectrum, rather than a single molecular species (see Fig. 4.3).
For complex antigens (such as bacterial proteins with multiple epitopes), the antibodies produced are different in chemical structure and specificity, because they are formed by different B cell clones, and each clone expresses a type that can interact with the antigen.
Different epitopes of immunoglobulins (Chapters 6 and 14). This makes the biochemical study of immunoglobulins very difficult, because a single specific pure molecule is not easy to separate. A contributing finding in this regard is that after serum electrophoresis, the immunoglobulin region (Chapter 35) of patients with B-cell malignancies with myeloma is usually a band with a very narrow electrophoretic mobility range (Fig 4.4).
Fig 4.4 | Gel electrophoresis of sera from different patients.
In this disease, a single clone of B cells may proliferate. These cells secrete the same immunoglobulins and accumulate in relatively high concentrations in the serum. These immunoglobulins have become relatively pure protein sources in biochemical research in the early stages of their structural studies.
3.2 Antibody structure
All antibodies have the same basic molecular structure (Fig 4.5). They are composed of a light (L) chain and a heavy (H) chain, and these terms refer to their relative molecular weight. The molecular weight of the light chain is about 25,000, and the molecular weight of the heavy chain is about 50,000 to 70,000. In a basic immunoglobulin molecule, two heavy chains and two light chains are connected by intermolecular disulfide bonds, as shown in Fig 4.5.
The five different types of human heavy chains have slightly different structures and are represented by lowercase Greek letters: µ (mu) for IgM, δ (delta) for IgD, γ (gamma) for IgG, ε (epsilon) for IgE, α ( alpha) represents IgA (Table4-1). There are two types of light chains, κ (Kappa) or λ (lambda). These two types of light chains are found in all immunoglobulins, but any kind of antibody contains only one type of light chain. Any IgG molecule is composed of exactly the same H chain and exactly the same L chain, forming the Y-type structure shown in Fig 4.5.
Fig 4.5 | Basic structure of antibody
The basic immunoglobulin has a molecular part with unique functions. Many biochemical studies have proved this. If the basic immunoglobulin molecule (IgG) is cleaved by protease hydrolysis, several fragments will be produced (Fig 4.6). For example, if papain is used to cleave IgG, two main types of fragments will be obtained. A fragment binds to an antigen and is called fragment antigen binding (Fab).
The other fragment is the crystallizable fragment (Fc); it does not bind to the antigen, but activates a molecular pathway of complement (Chapter 20). Fc has a variety of biological effect functions, such as the ability to bind to Fc receptors on macrophages and various other cells.
If pepsin is used to hydrolyze immunoglobulins, the two Fab fragments will remain connected (F[ab’]2), but the Fc fragment will be digested into small fragments, and the effector function will be lost. These findings indicate that the different molecular parts of the Ig molecule have different functions, one is responsible for binding antigens, and the other performs other biological effect functions.
Fig 4.6 | Proteolytic digestion of immunoglobulins
Further biochemical studies proved that the L and H chains can be divided into highly variable regions (VL and VH) and substantially constant regions (CL and CH). For example, if amino acid sequencing is performed on several different lambda chains from different immunoglobulins, there will be a fairly similar region at the N-terminus of the L chain, but there will also be a region of approximately 110 amino acid residues that can be observed There are great sequence differences between different lambda chains (Fig 4.7).
Fig 4.7 | Immunoglobulin amino acid sequence: variable and constant regions
The same is true for the H chain. The C region performs biological effect functions, such as binding complement protein, and the V region binds antigen. These variable regions are essential for responding to a large number of different antigen structures.
In addition, the three-dimensional structure determination showed that immunoglobulins are composed of folded, repetitive fragments called domains. The L chain is composed of a variable domain and a constant domain, and the H chain is composed of a variable domain and three or more constant domains. Each domain is approximately 110 amino acid residues long, and is connected to other domains through short segments of a longer polypeptide chain, as shown in Fig 4.8.
Fig 4.8 | Three-dimensional structure of immunoglobulin molecules
Other molecules of the immune system have similar folded polypeptide domains, which gave rise to the term: immunoglobulin superfamily to describe this group of related proteins.
3.3 Part of the characteristics and biological characteristics of immunoglobulins
Antibodies appear as soluble proteins in the circulation and can also be expressed on the surface of B cells. The main function of all antibodies is to bind antigens and inactivate pathogens. For example, by agglutinating bacteria, bringing them together, preventing them from entering the host cell. If bacteria are enveloped by antibodies, the possibility of them being phagocytes will increase. Antibodies can also activate complement (Chapter 20) and can trigger a lysis reaction that destroys the cells to which the antibody binds. These five types of antibodies have different functions, which are the result of structural differences (Fig 4.9).
Fig 4.9 | Biological characteristics of different types of immunoglobulins
IgM is the main antibody in the early stage of immune response. It is a pentameric structure composed of five H2L2 units. Each unit is similar to IgG, connected by a linking chain (J), and has 10 potential antigen binding sites; therefore , It is the most effective antibody for agglutinating bacteria and activating complement.
IgD mainly exists on the surface of B cells as a receptor molecule and participates in the activation of B cells.
IgG is the most common antibody molecule in serum (see Table 4-1). It survives intact in the serum for the longest time (ie, the longest half-life), and can pass through the placenta to provide maternal protection for newborns.
IgE was originally designed to prevent parasitic infections. The antigen binds to IgE coupled to the Fc receptors on mast cells and basophils, and triggers allergic reactions by activating mast cells and releasing mediators such as histamine (Chapter 27).
IgA is the main immunoglobulin in secretions such as saliva, breast milk and tears. It is also abundantly present in the mucosal epithelial cells of the respiratory tract, genitals and intestines. IgA in secretions (sIgA) consists of two IgA molecules, one J chain and one secretory component molecule. The secretory component protects the molecule from proteolytic enzymes and promotes its transfer across epithelial cells into the secretion.
As shown in Table 4-1, immunoglobulins interact with multiple cell types through various Fc receptors present on cells. This interaction can recruit cells (such as inflammatory macrophages), which can then secrete cytokines that protect the host, as part of the response to foreign antigens.
TABLE 4-1 | Some properties of human immunoglobulin
4.1 Active immunity
Vaccination or active immunization has greatly reduced the incidence of some infectious diseases, including smallpox and polio. The key principle (Chapter 2) is that if you have been exposed to a pathogen before, if you encounter the pathogen again, it will induce a protective response. Active immunization may involve antigens prepared using recombinant DNA technology; for example, hepatitis B surface antigen (HBsAg) has been very successful in protecting individuals from this infection. Hepatitis B virus (HBV) is the main cause of hepatitis and is associated with a relatively high morbidity and mortality rate. This kind of antibodies produced by vaccination can make more than 90% of vaccinated people develop resistance to infection. In some countries, hepatitis B vaccine is now vaccinated in infancy. In other countries, it is only available to specific groups, such as health care workers. HBV uses HBsAg to bind to receptors on liver cells. Once the virus binds to the cell, it can enter the cell. The vaccine induces antibodies against HBsAg, which prevent the interaction between HBsAg and its receptor (Fig 4.10).
Fig 4.10 | Anti-hepatitis B surface antigen (HBsAg) antibody can prevent hepatitis B virus from infecting liver cells
4.2 Passive immunity
During routine blood tests, a pregnant woman was found to be infected with hepatitis B virus. A blood test showed that her blood had a high level of virus, and the baby was at high risk of infection. In many countries, this type of vertical transmission is the most common way for infants to contract the virus and then become a lifetime infection. In the case, the baby was injected with hepatitis B immune globulin (HBIG) 6 hours after birth. Subsequently, she was vaccinated with hepatitis B vaccine, and testing at the age of 1 year confirmed that she was not infected.
HBIG is made from the plasma of donors who have been vaccinated with hepatitis B vaccine and have produced large amounts of antibodies against hepatitis B surface antigen (HBsAg). It is more than 70% effective in preventing the vertical transmission of hepatitis B, and it is also used for post-exposure prevention, such as needlestick injuries. This antibody preparation provides immediate protection against the virus without the need for the body to react (passive immunity). The protective effect is only effective when the antibody persists; the serum half-life is 21 days. Passive immunity is also used to prevent other infections and has been tested in Ebola virus infection.
4.3 Therapeutic antibody
A 78-year-old woman with mild dementia fainted at home and was sent to the emergency department. She usually takes digoxin and diuretics to treat atrial fibrillation and heart failure. She had sinus bradycardia 47 beats per minute, and the attending doctor wanted to know if this might be the result of the patient taking too many digoxin pills. Her blood digoxin level is 7.2nmol/L, which is much higher than the treatment range (1.2~2nmol/L). The diagnosis was sinus bradycardia caused by accidental digoxin overdose.
The half-life of digoxin is very long, 36 hours. Due to severe bradycardia, the patient’s doctor decided to provide her with a Fab fragment against digoxin. After 4 hours of administration, the pulse rate had returned to normal, and the patient’s symptoms had improved significantly.
Anti-Digoxin Fab fragments are produced by inoculating sheep with digoxin and carrier protein. The carrier protein is necessary because digoxin acts as a hapten. Once the sheep has developed a good antibody level, a large amount of blood samples will be taken, and then the immunoglobulin will be separated from the sheep serum, treated with papain to produce Fab fragments, and then injected into the patient. Anti-Digoxin Fab fragments are successful because they have two characteristics of antibodies. The affinity of Fab fragments for digoxin is higher than that of digoxin for its pump receptors, allowing Fab fragments to replace digoxin and reduce its effect on cardiotoxicity. FAB fragments are also very specific and will not bind to anything else in the body, which reduces their risk of side effects.
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