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Medical Immunology Chapter 5 Immunoglobulin zhang lining Immunology Institute Medical School Shandong University Antibody:a type of glycoprotein molecule, produced by B cells that bind antigens often with a high degree of specificity The basic structural unit of antibody is composed of two identical heavy chains and two identical light chains • Immunoglobulin( Ig): It refers to all globulins that possess the bioactivity of Ab or a similar structure to Ab • Therefore, all Abs belong to Igs, but not all Igs possess the functions of Abs -globulin Section I. Molecular Structure of Immunoglobulin I. Basic structure of Ig Four polypeptide chains 2 identical heavy chains 2 identical light chains N-terminal end The 4 polypeptide chains are joined by S-S bonds. inter-chain disulfide bonds (S-S) intra-chain disulfide bonds (S-S) Two terminal ends “N” terminal end “C” terminal end Two regions variable regions constant region C-terminal end 1.Heavy and Light chain: . Heavy chains (H) 450 ~550aa, 50—75KD . Light chains (L) 214 aa, 25KD Classes and types of Ig According to the differences of H chains( amino acid composition , sequence), H chains can be divided into 5 classes and their constituted Ig can be classified into 5 classes. • Five classes of H Chain: • Five classes of Ig : IgG IgA IgM IgD IgE According to the differences of L chains ( amino acid composition , sequence), L chains can be divided into 2 types and their constituted Igs can be typed into 2 types , and : 20:1 (in mice) 2:1 (in human) 2. variable regions and constant regions 1) Variable region (V): ½ of L chain or ¼ of H chain from N end 2) Constant region ( C ) ½ of L chain or 3/4 of H chain from C end (1) Variable region (V) Hypervariable region (HVR) Framework region (FR) Hypervariable region (HVR) Most of the sequence differences among antibodies are confined to three short stretches in the V regions of heavy and light chains are called HVR. Because these sequences form an antigen-binding surface that is complementary to the threedimensional structure of the bound antigen, HVR are also called Complementarity –determining regions, CDRs L chain possesses 3 CDRs:CDR1, CDR2 and CDR3 H Chain possesses 3 CDRs : CDR1, CDR2 and CDR3 • CDR: complementarity –determining regions The three short stretches in the V regions of Ig that contain most of the sequence differences among Igs are called CDR because these sequences form an antigen-binding surface that is complementary to the three-dimensional structure of the bound antigen L: CDR1—28-35, CDR2—49-56, CDR3—91-98 H: CDR1-29-31 CDR2-49-58 CDR3-95-102 CDR1, CDR2,CDR3 CDR1 CDR2 CDR3 Ag-binding sites L: CDR1—28-35, CDR2—49-56, CDR3—91-98 H: CDR1-29-31 CDR2-49-58 CDR3-95-102 The antigen-binding site binds to epitope of antigen CDR3 CDR1 CDR2 Ag-binding sites 3. Hinge region • The hinge region is segment of heavy chain between the CH1 and CH2 domains. • Flexibility in this area permits the two antigenbinding sites to operate independently. II. Other components of Ig • Joining chain(J ) Secretory piece( SP) Joining chain (J ) produced by plasma cells Functions: linker Join monomer of Ig to form dimer, or polymer ( Ig A, IgM) Secretory piece( SP) . produced by mucosa epithelial cells . Bind to dimer of IgA to form secretory IgA (SIg A) . Functions: protect SIgA against proteolysis in secretory liquid. Secretory piece( SP) J chain Secreted piece Formation of secreted piece III. Domains of Ig Domains : the Polypeptide chains of Ig are folded by intrachain s-s bond into globular shape in each 110aa regions which is called a domain • The domains of L chain: 2 domains ,VL and CL • The domains of L chain: 4-5 domains 4 domains: VH, CH1, CH2, CH3 in IgG, IgD, IgA 5 domains : VH, CH1, CH2, CH3 CH4 in IgM, IgE Function of domains • VH , VL: antigen-binding site • CH1, CL: genetic marker • 2CH2: complement-fixing site or crossing the placenta • CH3/CH4: cell-binding site hinge region : flexible and suitable for CDR of Ig bond to antigenic determinants. IV. Proteolytic fragments of an IgG molecules and their functions Ig digested by papain and pepsin .position .Fragments: .Function: 1. Products of IgG digested by papain • Position : near the N terminus of S-S bonds of H inter-chains fragments: 2Fab (antigen-binding fragment) Fc (crystalizable fragment) Function: Fab : bind specifically to Ag Fc 1) fix complement 2) cross the placenta 3) bind to FcR on different cells • 2. Products of IgG digested by pepsin • Position: near the S-S bond of H inter-chains from the C end • Fragments and function : F(ab’)2 : bind to antigen (2 values) pFc’ : no function Significance · Elucidating the relationships between the structure and function of Igs . Decrease the immunogenicity of Ig for clinical treatment Section II .The features and functions of different classes Ig I . IgG 1. Highest concentration in serum (75% of total Ig) 2. Four subclasses : IgG1, IgG2, IgG3, IgG4 3.Unique Ig that can pass through placenta. 4.half-life is longest one among all Igs ( 20-23 days ) 5. starts to be produced at 2-3 month after birth and reach the level of adult at 5 -years old 6. Functions of IgG: • Important Ig against bacteria and virus,neutralize toxin • Some IgG belong to the auto-antibodies: eg. long active thyroid stimulator (LATS) • combine with the Fc receptor(FcγR) •combine with the Fc receptor(FcγR) II. IgA 1.Two types serum type :monomer secretary type(sIgA): dimer,trimer or polymer 2.two subclasses:IgA1,IgA2 II. IgA 1.Two types serum type :monomer secretary type(sIgA): dimer,trimer or polymer 2.two subclasses:IgA1,IgA2 3. to be produced at 4 month after birth 4. activate the complement by alternative pathway 5.local immunity of mucosa • local immunity neutralize virus/toxin III. IgM 1. MW is highest :pentamer(90KD) Valences:10 valences in theory 2.half-life is shorter(4~5 days) 3.The earliest synthesized Ig • be produced during fetus • appear in the early stage after infection 4. The mIg of the B cells act as the antigen receptors of B cells(BCR) 5.Function: • IgM is more effect in anti-infection anti-bacterium • natural Ab for blood-type antigen • auto-antibody :rheumatoid factor(RF) IV. IgD 1. The concentration in serum is low and sensitive to proteinase 2. Act as the antigen receptor on B cells (mIgD): B cells Regulate the differentiation of V. IgE 1.Concerntration of IgE in serum is the lowest in normal individual, but is very high in some patients 2.related to type I hpersensitivity high affinity to FcR of mast cells and basophils • Section III Biological function of Ig I. The function of V region -binds to antigen specifically Neutralization II. The function of C region 1. Activate complement Complement lesion II. The function of C region 2. bind to Fc receptor on some cells (1)Opsonization(IgG,IgM) enhance the phagocytosis of MΦ II. The function of C region 2. bind to Fc receptor on some cells (2)ADCC( antibody dependent cell mediated cytoxicity II. The function of C region 2. bind to Fc receptor on some cells (3) Type I hypersensitivity -mast cell (FcR) - basophils (FcR) II. The function of C region 3. Cross placenta or mucosa The IgG from mother pass through the placenta into fetal body by binding of IgG with receptor, FcRn on placenta Section IV The Immunogeneity of Ig • isotype • allotype • idiotype I. Isotype of Ig The epitope of Ig existing in all healthy individuals of a species is called as isotype This is a kind of species specificity which exists in C region of immunoglobulin, including class, subclass, type, subtype The isotypes of Ig 1.Classes: Heavy chains: ,,,, Igs: IgG,IgM,IgA,IgD, IgE 2.subclasses: IgG: IgG1-4; IgA1-2 3. types: Light chains: , Igs: IgG type or type etc. 4. Subtypes: :OZ(+),OZ(-) II. Allotype: The property of a group of antibody molecules defined by their sharing a particular antigenic determinant found on the antibodies of some individuals but not others of a species This is a kind of individual specificity within a species which exists in C region of immunoglobulin III. Idiotype of Ig Igs produced by one B cell clone possess unique structure respectively in hypervariable region(HVR) ,this unique structure of Ig is called idiotype of Ig In fact: HVR CDR idiotype are in the same sites of Ig Section V. Preparation of Ab Polyclonal Ab Monoclonal Ab Gene engineering Ab I.Polyclonal Ab a mixture of Abs with different specificities and affinities generated in a natural response or artificial immunization II. Monoclonal Ab (mAb) Abs produced by single B cell clone (or one hybridoma clone ) possess same structure and specificity. mAb / McAb Prepared by hybridoma technique: Immunized spleen cells (B) fuse with myeloma cells and form hybridoma with property of proliferating and producing antibody III. Gene engineering Ab Abs prepared by the method of gene recombination Review for chapter 6 immunoglobulin Terms: Antibody , Immunoglobulin, CDR , domain of Ig ,Polyclonal Ab, Monoclonal Ab, genetic engineering Ab Key questions : .Describe the basic structure of Ig .Describe the composition and functions of each domain of IgG .Describe the functions and hydrolysis fragments of Ig digested by Papain /pepsin .Please expatiate the features and functions of five classes Ig .How to understand the the biological efforts of Ab New Thinking way To my students: Why not the best