The immune system has a series of dual natures, the most important of which is self/non-self recognition. There are two types of immunity—innate (non-specific) and specific (adaptive) immunity. The innate immunity system is what we are born with and it is non-specific (Table 7.1). All types of antigens are attacked pretty much equally present in all normal humans. It is genetically based and we pass it on to our offspring. The human body has various methods of defence which are as follows:
- The most important barrier is the skin. The skin cannot be penetrated by most organisms unless it already has an opening, such as a nick, scratch or cut.
- Mechanically, pathogens are expelled from the lungs by ciliary action as the tiny hairs move in an upward motion; coughing and sneezing abruptly eject both living and non-living things from the respiratory system; the flushing action of tears, saliva and urine also force out pathogens, as does the sloughing off of skin.
- Sticky mucus in respiratory and gastrointestinal tracts traps many microorganisms.
- Acid pH (<7.0) of skin secretions inhibits bacterial growth. Hair follicles secrete sebum that contains lactic acid and fatty acids both of which inhibit the growth of some pathogenic bacteria and fungi. Areas of the skin not covered with hair, such as the palms and soles of the feet, are most susceptible to fungal infections.
- Saliva, tears, nasal secretions and perspiration contain lysozyme, an enzyme that destroys Gram positive bacterial cell walls causing cell lysis. Vaginal secretions are also slightly acidic (after the onset of menses). Spermine and zinc in semen destroy some pathogens. Lactoperoxidase is a powerful enzyme found in mother's milk.
Table 7.1 Differences Between Non-specific and Specific Immune Responses
Non-specific Immunity Specific Immunity Antigen-independent immunity Antigen-dependent immunity Immediate maximum response Time dependent with a lag and memory based Non-antigen specific Antigen-specific Temporary Permanent, memory based
- The stomach is a formidable obstacle insofar as its mucosa secrete hydrochloric acid (0.9 < pH < 3.0, very acidic) and protein-digesting enzymes that kill many pathogens. The stomach can even destroy drugs and other chemicals.
A very important group of cells known as lymphocytes, which are widely distributed throughout the tissues, appear in increased number during infection resulting in inflammatory response which is followed by immune response. The introduction of an antigen or infection of the tissues by an invading organism results in increased proliferation of lymphocytes in the tissues of the reticulo-endothelial system particular the lymph nodes and the spleen. Subsequently, increase in the number of antibody-producing cells known as plasma cell can be seen.
There is an initial lag period following the detection of the antigen after which antibody may be demonstrated in the serum, its concentration increasing up to a maximum level before declining. An animal will show different responses if the antigen is completely new to it (a primary response) or if the antigen has been encountered on a previous occasion (a secondary response). Figure 7.1 shows primary and secondary response against the antigen. The second and subsequent injections are known as booster doses. The secondary response shows reduced lag period and a considerably increased rate of antibody synthesis compared with the primary response and the antibody persists for a longer period.
The cells together comprising as lymphocytes constitute the defence against an attack by an organism. The majority of lymphocytes are to be found in tissues collectively known as reticulo-endothelial system. This includes such tissues as liver, spleen, bone marrow, thymus and lymph nodes. All these organs are sources for producing immune responses.
Figure 7.1 Production of Antibodies in Response to Antigen Injected
- Cell-mediated immunity is because of a sub-population of T-lymphocytes. These lymphocytes are derived from thymus.
- Humoral immunity is because of sub-population of B-lymphocytes. These lymphocytes originate in men in the bone marrow and B-cells are antibody-producing cells.
The introduction or encounter of an antigen in the circulating fluid results in increased production of lymphocytes particularly by lymph nodes and spleen. Subsequently, antibody producing lymphocytes can be observed. The raising of a specific antiserum by the immunization of an animal usually involves the parenteral injection of the pure antigen often introduced intramuscularly, although intravenous injection may be appropriate for particulate antigens. The feature of the primary and secondary responses to antigens suggests that a series of injections involving small amount of antigen are likely to be more effective than a single large injection. Their precise sequence and timing can significantly affect the quality of the antiserum produced. An initial injection is normally followed by several booster doses given at 2 to 4 week intervals. Too frequent injection, although possibly giving a quicker response, may result in an antiserum of reduced avidity.
The species of animals used should be as different as possible from the animal which is the source of the antigen. It should be relatively easy to handle and yet provide enough serum to make the process worthwhile. The animals most frequently used for the production of antibodies against human antigens are the guinea pig and rabbit, but for larger supplies of serum, goats and horses may be used.
Antigens vary considerably in their ability to initiate an immune response, and it is usual to incorporate an adjuvant into the sample prior to injection. An adjuvant is a mixture of substances which stimulates an inflammatory response and prevents the rapid removal of the antigen from the tissues by the normal drainage mechanism. Freund's adjuvant consists of an emulsion of dead mycobacteria in mineral oil but simpler alternatives of aluminium phosphate or hydroxide have a similar effect. Of all the naturally occurring substances, proteins are generally the most immunogenic and need to have a relative molecular mass of at least 4,000 and some structural rigidity to be effective as antigen and able to elicit antibody response. In order to raise antibodies against a non-immunogenic molecule (called hapten), it is necessary to link it to a carrier protein which is capable of initiating a response. Bovine or human serum albumin is frequently used for this purpose as well as synthetic polypeptides such as poly-L-lysine. The hapten should be linked covalently with the carrier protein, a process usually achieved by a fairly simple reaction using a carbodiimide.
Immunoglobulins (Ig) are glycoprotein molecules that are produced by plasma cells in response to an immunogen (antigen) and these immonoglobulins function as antibodies. The antibodies are also called immunoglobulins because they migrate with globular proteins when an antibody-containing serum is placed in an electrical field (Figure 7.2).
Figure 7.2 Electrophoretic Pattern of Human Serum
The basic structure of an immunoglobulin is illustrated in Figure 7.3. Although different immunoglobulins can differ structurally, they are all built from the same basic units. All immunoglobulins have a four chain structure as their basic unit. They are composed of two identical light chains (23 kD) and two identical heavy chains (50–70 kD).
The two light and the two heavy chains are held together by inter-chain disulfide bonds and by non-covalent interactions. The number of inter-chain disulfide bonds varies among different immunoglobulin molecules. Within each of the polypeptide chains there are also intra-chain disulfide bonds.
When the amino acid sequences of many different heavy and light chains are compared, it becomes clear that both the heavy and light chain could be divided into two regions based on variability in the amino acid sequences. These are the:
Figure 7.3 Schematic Diagram of Immunoglobulin G (IgG) with Two Heavy and Light Chains Connected by –S–S– Bridges
- Light chain—VL (110 amino acids) and CL (118 amino acids)
- Heavy chain—VH (110 amino acids) and CH (330–440 amino acids)
Each chain contains a variable region. The variable region has a specific hyper-variable region (HYV) which determines specificity and complementarity of the antibody to antigen.
The position at which antibody molecule forms a Y is called the hinge region. It is called the hinge region because there is some flexibility in the molecule at this point.
Three-dimensional images of the immunoglobulin molecule show that it is not straight as depicted; rather, it is folded into globular regions each of which contains an intra-chain disulfide bond. These regions are called domains.
Carbohydrates are attached to the CH2 domain in most immunoglobulins. However, in some cases carbohydrates may also be attached at other locations.
Comparisons of the amino acid sequences of the variable regions of immunoglobulins show that most of the variability is present in regions called the hypervariable regions (HYV) or the complementarity-determining regions (CDR) as illustrated in Figure 7.3. Antibodies with different specificities (i.e. different combining sites) have different CDR while antibodies of the exact same specificity have identical complementarity-determining region. CDR is the antibody combining site. The hypervariability determines the specificity of antibody. Thus, antibodies produced in response to particular antigen are specific for that antigen only.
The regions between the CDR in the variable region are called the framework regions. Based on similarities and differences in the framework regions, the immunoglobulin heavy and light chain variable regions can be divided into groups and subgroups. These represent the products of different variable region genes.
Immunoglobulin fragments produced by proteolytic digestion have proven very useful in elucidating structure–function relationships in immunoglobulins.
Digestion with papain breaks the immunoglobulin molecule in the hinge region before the H-H inter-chain disulfide bond (Figure 7.3). This results in the formation of two identical fragments that contain the light chain and the VH and CH1 domains of the heavy chain.
Antigen Binding: These fragments were called the Fab fragments because they contained the antigen binding sites of the antibody. Each Fab fragment is monovalent whereas the original molecule was divalent. The combining site of the antibody is created by both VH and VL. An antibody is able to bind a particular antigenic determinant because it has a particular combination of VH and VL. Different combinations of a VH and VL result in antibodies that can bind a different antigenic determinant.
Digestion with papain also produces a fragment that contains the remainder of the two heavy chains each containing a CH2 and CH3 domain. This fragment was called Fc because it was easily crystallized.
Effector Functions: The effector functions of immunoglobulins are mediated by this part of the molecule. Different functions are mediated by the different domains in this fragment (Figure 7.3). Normally the ability of an antibody to carry out an effector function requires the prior binding of an antigen; however, there are exceptions to this rule.
Treatment of immunoglobulins with pepsin results in cleavage of the heavy chain after the H-H inter-chain disulfide bonds resulting in a fragment that contains both antigen binding sites (Figure 7.3). This fragment was called F(ab′)2 because it is divalent. The Fc region of the molecule is digested into small peptides by pepsin. The F(ab′)2 binds antigen but it does not mediate the effector functions of antibodies.
Table 7.2 Classes of Immunoglobulins
The immunoglobulins can be divided into five different classes, based on differences in the amino acid sequences in the constant region of the heavy chains (Table 7.2). All immunoglobulins within a given class will have very similar heavy chain constant regions. These differences can be detected by sequence studies or more commonly by serological means (i.e. by the use of antibodies directed to these differences).
Immunoglobulins can also be classified based on the type of light chain they have. Light chain types are based on differences in the amino acid sequence in the constant region of the light chain. These differences are detected by serological means.
- Kappa light chains
- Lambda light chains
IgG: It comprises some 80 per cent of the total immunoglobulin in plasma and because it is relatively small. It is capable of crossing membranes and diffusing into the extravascular body spaces. It can cross the placental membrane and provides the major immune defence during the first few weeks of life until the infant's own immune mechanism becomes effective.
IgM: It is a large molecule composed of five units, each one similar in structure to an IgG molecule. The tetramer contains an additional polypeptide, the J chain (relative molecular mass 15,000) which appears to be important in the secretion of the molecule from the cell. It is an effective agglutinating and precipitating agent and, although potentially capable of binding ten antigen molecules, it is usually only pentavalent. It does not cross membranes easily and is largely restricted to the bloodstream.
IgA: It is associated mainly with seromucous secretions such as saliva, tears, nasal fluids, etc., and is secreted as a dimer with both a J chain and a secretor piece (relative molecular mass 70,000), the latter apparently to prevent damage to the molecule by proteolytic enzymes. Its major role appears to be the protection of mucous membranes and its presence in blood, mainly as the monomer, may be as a result of absorption of the degraded dimer.
IgE: It is known as a cytophilic immunoglobulin because of its ability to bind to cells which may account for its low concentration in body fluids. When IgE reacts with an antigen it causes degranulation of the mast cell to which it is bound with the release of vasoactive amines such as histamine. This process may well be helpful in initiating the inflammatory response but in allergic individuals the reaction is excessive and leads to a hypersensitive or over-reactive state.
MHC is a group of genes on a single chromosome (chromosome 6p21.3, in humans) that codes the MHC antigens. Human leukocyte antigens (HLA) are the MHC antigens of humans, and called so because they were first detected on leukocytes. A set of MHC alleles present on each chromosome is called an MHC haplotype.
The Human Leukocyte Antigens (HLA) system contributes to the immune response. It is a set of molecules (glycoproteins) expressed at the surface of almost all cells that are responsible for lymphocyte recognition and ‘antigen presentation’. The HLA molecules control the immune response through recognition of ‘self’ and ‘non-self’. They belong to a group of molecules known as the immunoglobulin supergene family, which includes immunoglobulins, T-cell receptors, CD4, CD8 and others. The main function of the HLA molecules is presenting the antigen (protein chain of antigen) to the T Lymphocytes and initiating the specific immune response.
HLA molecules are coded by two groups of genes, HLA class I and HLA class II, and the functions of both groups are really quite distinct (Figure 7.4).
HLA Class I Proteins
They are coded by the genes HLA-A, HLA-B and HLA-Cw. Class I molecules are found on virtually every cell in the human body and they present antigen to cytotoxic T-cells (CTLs) (the CD8 + T Cell). Class I molecules present ‘endogenous’ antigen. An endogenous antigen might be fragments of viral proteins or tumour proteins. Presentation of such antigens would indicate internal cellular alterations that if not contained could spread throughout the body. Hence, destruction of these cells by CTLs is advantageous to the body as a whole. Class I molecules are made up of two chains, a heavy chain (transmembrane polypeptide) coded by the genes HLA-A, HLA-B and HLA-Cw, and a light chain β-2 microglobulin (a non-transmembrane polypeptide).
HLA Class II Proteins
They are coded by the gene HLA-DR, HLA-DQ and HLA-DP. Class II molecules, in contrast to Class I molecules, are found only in B-cells, macrophages and other ‘antigen-presenting cells’ (APCs). Class II molecules present antigen to helper T-cells (TH-cells) (CD4 + T cells—The CD4 + T cells that activate B cells are called Helper T cells.). Class II molecules present ‘exogenous’ antigens. Exogenous antigens, in contrast, might be fragments of bacterial cells or viruses that are engulfed and processed by, for example, a macrophage and then presented to helper T-cells. The TH-cells, in turn, could activate B-cells to produce antibody that would lead to the destruction of the pathogen. Class II molecules consist of two transmembrane polypeptides, α chain and β chain. The β chain is much more polymorphic compared to the α chain. For this reason, HLA typing is currently done on β chain (HLA-DRB1, HLA-DRB3, HLA-DRB4, HLA-DRB5, HLA-DQB1 and HLADPB1).
Figure 7.4 MHC of Humans at Chromosome 6p21.3
Class III region is not actually a part of the HLA complex, but is located within the HLA region, because its components are either related to the functions of HLA antigens or are under similar control mechanisms to the HLA genes. Class III antigens are associated with proteins in serum and other body fluids (e.g. C4, C2, factor B, TNF) and have no role in graft rejection.
Histocompatibility genes are inherited as a group (haplotype), one from each parent. Thus, MHC genes are co-dominantly expressed in each individual. A heterozygous human inherits one paternal and one maternal haplotype, each containing three Class-I (B, C and A) and three Class II (DP, DQ and DR) loci. Each individual inherits a maximum of two alleles for each locus. The maximum number of class I MHC gene products expressed in an individual is six; that for class II MHC products can exceed six but is also limited. Thus, as each chromosome is found twice (diploid) in each individual, a normal tissue type of an individual will involve 12 HLA antigens. Haplotypes, normally, are inherited intact and hence antigens encoded by different loci are inherited together. However, on occasions, there is crossing over between two parental chromosomes.
Because some HLA antigens are present on almost all of the tissues of the body, the identification of HLA antigens is also described as ‘tissue typing’. HLA matching between donor and recipient is desirable for allogenic transplantation.
For haemopoietic stem cell (HSC) transplants, the degree of HLA matching is critical in determining the probability of Graft-versus-Host Disease (GvHD). This is the major form of rejection in clinical HSC transplants. In an attempt to minimize these allo-responses, the HLA class I and class II types of the donor and recipient are matched as closely as possible. However, because of extensive polymorphism, an HLA identical donor is only rarely available. Most transplant recipients therefore receive immunosuppressive drugs to prevent or stop detrimental allo-responses, but this non-specific approach also compromises beneficial immune responses to infection. Most centres will only transplant across 1 HLA class I (HLA-A or B locus) difference.
The actual HLA testing is performed on a blood sample from the patient and potential donors. There are different ways that an HLA typing test can be done. These methods differ in their ability to detect differences between a patient and donor. HLA typing can be performed by serologic typing or by DNA molecular typing techniques. Serologic typing is often the first step in HLA typing, which identifies the major transplantation antigens that make up a patient's HLA (at least A, B and DR, but now matches are defined to a level of 10/10 on loci A, B, Cw, DR and DQ). This test can be done rather quickly with results available within 2 to 3 days. Although serologic testing is sufficient in some cases to identify a compatible sibling donor, more sensitive techniques using molecular typing of DNA are needed for identification of compatible ‘unrelated’ donors (a first quick search is done using serological or molecular low-resolution methods, and a second more specific search is carried out using high-resolution methods on locus A,B, Cw, DR and DQ and facultative DP).
Class I typing methods include test such as microcytotoxicity (for typing A, B, C loci) and cellular techniques such as CML (for HLA-DPw typing). Class II typing involves cellular techniques such as mixed leukocyte reaction (MLR)/mixed leukocyte culture (MLC) (for DR typing) and molecular techniques such as PCR and direct sequencing (for DR, DQ typing).
Serological Tissue Typing Method
Serologic-based HLA typing uses antigen-specific sera to determine a person's HLA type. Sera are human-derived preparations that react to specific HLA antigens expressed on white blood cells. A serologic-based test determines a person's HLA tissue type by noting which types of sera react to the person's white blood cells and which sera do not react. After an incubation step that permits time for the antibody to bind to any corresponding antigens in the cells, a complement is added to facilitate cell lysis. The reactions are then viewed under the microscope and graded according to the amount of cell lysis that has occurred. The type is assigned after reviewing the reaction patterns for the various sera. Serologic-based typing is a less specific test than molecular-based tissue typing.
Disease Associated with Class I HLA: Ankylosing spondylitis (B27), Reiter's disease (B27), Acute anterior Uvietis (B27), Psoriasis vulgaris (Cw6).
Disease Associated with Class II HLA: Hashimoto's disease (DR5), Primary myxedema (DR3), Graves thyrotoxicosis (DR3), Insulin-dependent diabetes (DQ2/8), Addison's disease (adrenal) (DR3), Goodpasture's syndrome (DR2), Rheumatoid arthritis (DR4), Juvenile rheumatoid arthritis (DR8), Sjogren's syndrome (DR3), Chronic active hepatitis (DR3), Multiple sclerosis (DR2, DR6), Celiac disease (DR3), Dermatitis herpetiformis (DR3) No definite reason is known for this association. However, several hypotheses have been proposed: antigenic similarity between pathogens and MHC, antigenic hypo- and hyper-responsiveness controlled by the class II genes are included among them. Possible explanation for these associations is that the HLA antigen itself plays a role in disease, by a method similar to one of the following models.
Oxygen is the terminal acceptor of electrons in all aerobically living organisms, including humans. The oxidation of energy rich compounds such as NADH and FADH2 in mitochondria during energy generation process is a fundamental process and oxygen is converted to water at the end of the respiratory chain in the mitochondria. However, in the same mitochondrial respiratory chain, incomplete reduction of oxygen leads to generation of superoxide. Superoxide is a radical, that is, a chemical species with an unpaired electron. Radicals, with lone pair of electrons, such as superoxide anion, are very reactive species and initiate further ‘cascade’ of chain reactions generation other reactive species. The superoxide radical, hydroxyl radicals are known as ‘reactive oxygen species’ (ROS) which bring about damage to organs. This process is known as oxidative damage due to ROS or simply oxidative stress. The production of superoxide by the mitochondrial respiratory chain occurs continuously during normal aerobic metabolism. It has been estimated that 1 to 2 per cent of all the electrons travelling down the mitochondrial respiratory chain never make it to the end, but instead form superoxide. In addition to the mitochondrial respiratory chain, there are other endogenous sources of superoxide production.
In the presence of metal ion such as iron, in tissue like brains H2O2 though not a free radical, but is rapidly decomposed via the Fenton reaction generating hydroxyl radical:
In addition, superoxide, hydrogen peroxide and hydroxyl radicals can be interconverted via the so-called Haber–Weiss reaction:
Cuprous and cupric ions may substitute for ferrous and ferric ions in the Haber–Weiss reaction. Peroxy-nitrite can be formed from the reaction of NO with superoxide:
The ROS formed by the mechanisms explained above can cause oxidative damage to various biological molecules. Oxidative stress generally describes a condition in which cellular antioxidant defences are inadequate to completely detoxify the free radicals being generated, due to excessive production of ROS, loss of antioxidant defences or, typically, both. This condition may occur locally, as antioxidant defenses may become overwhelmed at certain subcellular locations while remaining intact overall, and selectively with regard to radical species, as antioxidant defences are radical-specific, for example, SOD for superoxide and catalase or glutathione peroxidase for H2O2.
Hydroxyl radicals can damage cell membranes and lipoproteins (the particles carrying cholesterol and fat in the blood stream) by a process called lipid peroxidation (LPO). The LPO can be measured by measuring melanoaldehyde (MDA) Lipid peroxidation occurs by a radical chain reaction, that is, once started, it spreads rapidly and affects a great number of lipid molecules. Proteins may also be damaged by ROS, leading to structural changes and loss of enzyme activity. (An enzyme is a protein that catalyzes the rate of a chemical reaction without itself being changed.)
The reversible nature of the antigen–antibody reaction has facilitated the development of competitive binding techniques with wide-ranging applications and high levels of sensitivity and specificity. The basic concepts of these techniques lie in the competition for a limited amount of antibody between the test antigen and a constant amount of reference antigen. The reference antigen, although identical immunologically, is distinguishable from the test antigen by the incorporation of an easily detectable group, known as the ‘label’. The nature of the label used provides a means of classifying the various techniques, for example, enzyme-linked immunoassay.
When two types of antigen are present, labelled reference (Ag*) and unlabelled test (Ag), the basic reaction may be represented by the following equation:
Provided that only a limited amount of antibody (Ab) is used, the equilibrium mixture will contain all of these components except free antibody and may be represented as follows:
In an analytical method the amount of antibody and labelled antigen are held constant and the only variable in the system is the amount of the test antigen. As the amount of this component varies, so the relative proportions of the other components in the equilibrium mixture will vary. An increase in the amount of the test antigen will result in an increased amount binding to the antibody (AgAb). This will force a parallel decease in the labelled antigen bound to antibody (Ag*Ab). Hence, the amount of labelled antigen bound to antibody in the resulting equilibrium mixture will be inversely proportional to the amount of test antigen introduced into the assay. Under these conditions, the amount of test Ag is
Such a relationship forms the quantitative basis of competitive binding immunoassays and also highlights the two major technical aspects of such techniques.
The name ELISA is an acronym coined by the developers of the method and stands for enzyme-linked immunosorbent assay. It was developed primarily to ease the process by which the bound antigen is separated from the free and also to use a label other than a radioisotope.
The method involves the binding of an antibody to the walls of a plastic tube and subsequently adding the test and labelled antigens which compete for the available antibody. After a suitable reaction time, the tube is washed to remove all the reagents except those which are bound to the antibody. The amount of enzyme-labelled antigen which is bound to the tube wall can be measured by monitoring the enzymic reaction after the addition of a suitable substrate. Under carefully controlled conditions, the amount of product formed is proportional to the amount of enzyme present and inversely proportional to the amount of test antigen.
The method is probably more useful in what is known as double antibody or sandwich form which uses enzyme-labelled antibody rather than antigen. In this form, the method is not competitive in nature and the quantitative aspects become more direct as a result. The specific antibody is bound to the walls of the vessel but instead of using a labelled antigen, it is added and allowed to react. After washing the tube well to remove all the other components, an enzyme conjugate of the same specific antibody is added which binds to the antigen molecules already held by the solid-phase antibody. Conjugate is washed away and a suitable substrate added and the amount of product formed in a given time is directly proportional to the amount of antigen originally present.
Myosin is one of the most abundant proteins in the human body. It is found in all the body's muscle types, in the ears and eyes, in the blood platelets, and is used in cytokinesis. Because of all the diverse functions of myosin, it can be grouped into anywhere from seven to fourteen unique categories.
The most common type of myosin is myosin class II. This is the type present in muscle tissues. Class II myosin is used to contract muscle tissue, thereby giving an organism mobility. Myosin II is a component of the myofibres in skeletal, smooth and cardiac tissue. Each muscle in the body is composed of bundles of muscle fibres. These muscle fibres in turn are made up of many myofibrils which are components of both thin and thick myofilaments. The thin myofilaments are primarily made of actin protein while myosin marks the foundation for the thick myofilaments. These myosin myofilaments are in turn made of many overlapping myosin II proteins (Figure 7.5).
Figure 7.5 Structure of Myosin Protein
The myosin filaments lie next to the actin filaments and have the capability to temporarily bind to the actin, causing the muscle to move. This binding capability provided the basis of the sliding filament model of muscle contraction. This model proposes the myosin head binds to the actin filament and then rotates to a different position, possibly as much as a 45 degree change, which can be accomplished by a change in structure. The myosin head is comparable to a stretched spring held in place by free energy generated by ATP hydrolysis.
The myosin II protein is able to perform functionally and enzymatically as ATPase because of its dimeric shape. Myosin is a hexamer with a total molecular weight of 520 kD. It is composed of two heavy chains and four light chains. The two heavy chains each weigh 220 kD and begin with a globular head at the N-terminal and end with an α-helix at the C-terminal. The tail (C-terminal) region is periodically interspersed with hydrophobic residues to give a ‘coiled coil’ type rod. The amino acids in the C-terminal are non-helical which aids in stabilizing the myosin filament backbone. Furthermore, the backbone contains inter-myosin ionic bonds to assist in stabilization. The tails are connected to the heads at the neck, which is the location of the hinge area.
The four light chains weigh about 20 kD each and are paired into two regulatory light chains and two essential light chains. Each head of the protein has one chain of each type, giving each head a pear type shape.
The primary function of the human lens is to focus light undistorted onto the retina. While the transmission properties of most of the components of the eye are stable, the transmission properties of the lens change throughout life.
The structure of the human lens is seen in Figure 7.6. The lens is a transparent organ located behind the cornea and the iris. The outer edge of the lens consists of a single layer of epithelial cells, and a membrane that covers the entire organ. Lens epithelial cells do not divide except when undergoing repair. Some epithelial cells lose their nuclei and other organelles and become lens fibre cells. These lens fibre cells are filled with a 30 per cent solution of protein, known as cytosol (soluble) lens protein. Because there is little protein turnover in the lens fibre cells, damage to lens protein accumulates throughout life.
Figure 7.6 Structure of Human Eye Lens
- Discuss the structure of IgG. What are the regions in IgG responsible for specificity?
- Classify immunoglobulins.
- Write short notes on:
- Non-specific immunity
- What is HLA? How is HLA responsible for tissue specificity?
- What is the basis of electrophoresis? How is ELISA used for quantification of immunoglobulins?
Multiple Choice Questions
- The B lymphocytes are responsible for which immunity.
- Cell mediated
- The basic unit of all immunoglobulin molecules consists of how many chains which are linked by disulfide bonds.
- Immunoglobulins composed of more than one basic monomeric unit are termed as
- IgM are
- The part of the antibody molecule which combines with antigens is formed by a few amino acids in the F region of H and
- J chain
- H chain
- L chain
- M chain
- There are two major types of L chains in man, the kappa and
- Lamda chain
- Alfa chain
- Beta chain
- Gamma chain
- IgG comprises 80 per cent of the gamma globulins and contains 2-4 percent of
- Nucleic acids
- IgA has 5 to 10 per cent carbohydrate and does not cross the
- IgM is the major immunoglobulin expressed on the surface of which cells
- IgE is present in the serum in what concentrations
- Very High
- Very Low
- No antibody activity is associated with
- IgA are