免疫球蛋白的结构与功能的关系.pptx

Signalling antigen receptors on B cells-bifunctional antigen-binding secreted molecules（B 细胞表面受体和分泌的抗体）Structural conservation and infinite variability-domain structure（结构上不仅保守而且无限可变的）.The Immunoglobulin Gene Superfamily（免疫球蛋白的超家族）The immunoglobulin fold（免疫球蛋白的折叠）Framework and complementarity determining regions-hypervariable loops（框架结构和可变区）Modes of interactions with antigens（与抗原相互作用的模型）Effector mechanisms and isotype role of the Fc.（Fc 区的作用）Multimeric antibodies and multimerisationCharacteristics and properties of each Ig isotypeIg receptors and their functionsImmunoglobulin Structure-Function RelationshipCell surface antigen receptor on B cells B 细胞表面受体和分泌的抗体Allows B cells to sense their antigenic environmentConnects extracellular space with intracellular signalling machinerySecreted antibody（抗体）Neutralisation（中和作用）Arming/recruiting effector cells（激活或者诱导功能细胞）Complement fixation（帮助机体对抗原的清除）Immunoglobulin Structure-Function RelationshipImmunoglobulins are Bifunctional ProteinsImmunoglobulins must interact with a small number of specialised molecules-（免疫球蛋白必须与特殊分子相互作用）Fc receptors on cells（细胞表面的Fc受体）Complement proteins（辅助蛋白）Intracellular cell signalling molecules（细胞内信号转导分子）whilst simultaneously recognising an infinite array of antigenic determinants.（同时能够识别无限抗原族）Structural conservation and a capacity for infinite variability in a single molecule is provided by a DOMAIN structure.（结构上不仅保守而且无限可变的-抗体结构域）Ig domains are derived from a single ancestral gene that has duplicated,diversified and been modified to endow an assortment of functional qualities on a common basic structure（Ig结构域源于一个原始基因，复制，多元化，修饰等）Ig domains are not restricted to immunoglobulins（Ig 结构域不仅仅局限于免疫球蛋白）.The most striking characteristic of the Ig domain is a disulphide bond-linked structure of 110 amino acids long（Ig结构域最明显的特点是其双硫键，连接了110个氨基酸）.Immunoglobulin domainsThe genes encoding Ig domains are not restricted to Ig genes.Although first discovered in immunoglobulins,they are found in a superfamily of related genes,particularly those encoding proteins crucial to cell-cell interactions and molecular recognition systems.IgSF molecules are found in most cell types and are present across taxonomic boundariesIg gene superfamily-IgSFAntibodies are Proteins that Recognize Specific Antigens 抗体能够特异性的识别抗原抗体能够特异性的识别抗原Epitopes（抗原决定簇（抗原决定簇）:Antigen Regions that Interact with AntibodiesConsequences of Antibody Binding抗体结合效应抗体结合效应CLVLSSSSSSSSCH3CH2CH1VHFcFabF(ab)2Domains are folded,compact,protease resistant structuresDomain Structure of Immunoglobulins免疫球蛋白的结构域免疫球蛋白的结构域Pepsin cleavage sites -1 x(Fab)2&1 x FcPapain cleavage sites -2 x Fab 1 x FcLight chain Cdomainsk or lHeavy chain Cdomainsa,d,e,g,or mCH3CH3CH2CH3CH2CH1CH3CH2CH1VH1CH3CH2CH1VH1VLCH3CH2CH1VH1CLVLCH3CH2CH1VH1CLVLHingeCH3CH2CH1VH1VLCLElbowCH3CH2FbFvFvFvFbFvHingeElbowCH3CH2FbFvFlexibility andmotion of immunoglobulinsHingeFvFbFabCH3CH2CH1VH1VLCLFcElbowCarbohydrateThe Immunoglobulin FoldThe characteristic structural motif of all Ig domainsBarrel under constructionA barrel made of a sheet of staves arranged in a folded over sheetA b barrel of 7(CL)or 8(VL)polypeptide strands connected by loops and arranged to enclose a hydrophobic interiorSingle VL domainUnfolded VL region showing 8 antiparallel b-pleated sheets connected by loops.NH2COOHS SThe Immunoglobulin FoldImmunoglobulins must interact with a finite number of specialised molecules-Easily explained by a common Fc region irrespective of specificity-whilst simultaneously recognising an infinite array of antigenic determinants.In immunoglobulins,what is the structural basis for the infinite diversity needed to match the antigenic universe?Immunoglobulins are Bifunctional ProteinsAmino acid No.Variability8010060402020406080100120Cytochromes CVariability of amino acids in related proteinsWu&Kabat 1970Amino acid No.Variability8010060402020406080100120HumanIg heavychainsFR1FR2FR3FR4CDR2CDR3CDR1Distinct regions of high variability and conservation led to the concept of a FRAMEWORK(FR),on which hypervariable regions were suspended.Framework and Hypervariable regionsAmino acid No.Variability8010060402020406080100120Most hypervariable regions coincided with antigen contact points-the COMPLEMENTARITY DETERMINING REGIONS(CDRs)Hypervariable regionsHypervariable CDRs are locatedon loops at the end of the Fv regionsSpace-filling model of(Fab)2,viewed from above,illustrating the surface location of CDR loops Light chainsGreen and brownHeavy chainsCyan and blueCDRsYellowThe framework supports the hypervariable loopsThe framework forms a compact b barrel/sandwich with a hydrophobic coreThe hypervariable loops join,and are more flexible than,the b strandsThe sequences of the hypervariable loops are highly variable amongst antibodies of different specificitiesThe variable sequences of the hypervariable loops influences the shape,hydrophobicity and charge at the tip of the antibodyVariable amino acid sequence in the hypervariable loops accounts for the diversity of antigens that can be recognised by a repertoire of antibodiesHypervariable loops and framework:SummaryAntigens vary in size and complexityProtein:Influenza haemagglutininHapten:5-(para-nitrophenyl phosphonate)-pentanoic acid.Antibodies interact with antigens in a variety of waysAntigen inserts into a pocket in the antibodyAntigen interacts with an extended antibody surface or a groove in the antibody surfaceCH3CH2FbFvFvFvFbFvHingeElbowCH3CH2FbFvFlexibility andmotion of immunoglobulins30 strongly neutralising McAb60 strongly neutralising McAb Fab regions60 weakly neutralising McAb Fab regionsHuman Rhinovirus 14-a common cold virus30nmModels of Human Rhinovirus 14 neutralised by monoclonal antibodiesElectron micrographs of Antibodies and complement opsonising Epstein Barr Virus(EBV)Negatively stained EBVEBV coated with a corona ofanti-EBV antibodiesEBV coated with antibodies and activated complement componentsAntibody+complement-mediated damage to E.coliHealthy E.coliElectron micrographs of the effect of antibodies and complement upon bacteriaNon-covalent forces inantibody-antigen interactionsElectrostatic forcesAttraction between opposite chargesHydrogen bondsHydrogens shared between electronegative atomsVan der Waals forces Fluctuations in electron clouds around molecules oppositely polarise neighbouring atomsHydrophobic forcesHydrophobic groups pack together to exclude water(involves Van der Waals forces)Why do antibodies need an Fc region?Detect antigenPrecipitate antigenBlock the active sites of toxins or pathogen-associated moleculesBlock interactions between host and pathogen-associated moleculesThe(Fab)2 fragment can-Inflammatory and effector functions associated with cellsInflammatory and effector functions of complementThe trafficking of antigens into the antigen processing pathwaysbut can not activateStructure and function of the Fc regionCH3CH2IgA IgD IgGCH4CH3CH2IgE IgMThe hinge region is replaced by an additional Ig domainFc structure is common to all specificities of antibody within an ISOTYPE(although there are allotypes)The structure acts as a receptor for complement proteins and a ligand for cellular binding sitesMonomeric IgMIgM only exists as a monomer on the surface of B cellsCm4 contains the transmembrane and cytoplasmic regions.These are removed by RNA splicing to produce secreted IgMMonomeric IgM has a very low affinity for antigenCm4Cm3Cm2Cm1N.B.Only constant heavy chain domains are shownCm3 binds C1q to initiate activation of the classical complement pathwayCm1 binds C3b to facilitate uptake of opsonised antigens by macrophagesCm4 mediates multimerisation(Cm3 may also be involved)Cm4Cm3Cm2Cm1N.B.Only constant heavy chain domains are shownPolymeric IgMIgM forms pentamers and hexamersCCCCCCMultimerisation of IgMCm4Cm3Cm2CCCm4Cm3Cm2CCCm4Cm3Cm2CCCm4Cm3Cm2CCCm4Cm3Cm2CCs ss ss sCCs s1.Two IgM monomers in the ER(Fc regions only shown)2.Cysteines in the J chain form disulphide bonds with cysteines from each monomer to form a dimer3.A J chain detaches leaving the dimer disulphide bonded.4.A J chain captures another IgM monomer and joins it to the dimer.5.The cycle is repeated twice more6.The J chain remains attached to the IgM pentamer.Antigen-induced conformational changes in IgMPlanar or Starfish conformation found in solution.Does not fix complementStaple or crab conformation of IgMConformation change induced by binding to antigen.Efficient at fixing complementIgM facts and figuresHeavy chain:m-MuHalf-life:5 to 10 days%of Ig in serum:10Serum level(mgml-1):0.25-3.1Complement activation:+by classical pathwayInteractions with cells:Phagocytes via C3b receptorsEpithelial cells via polymeric Ig receptorTransplacental transfer:NoAffinity for antigen:Monomeric IgM-low affinity-valency of 2Pentameric IgM-high avidity-valency of 10IgD facts and figuresIgD is co-expressed with IgM on B cells due to differential RNA splicingLevel of expression exceeds IgM on nave B cellsIgD plasma cells are found in the nasal mucosa-however the function of IgD in host defence is unknown-knockout mice inconclusiveLigation of IgD with antigen can activate,delete or anergise B cellsExtended hinge region confers susceptibility to proteolytic degradationHeavy chain:d-DeltaHalf-life:2 to 8 days%of Ig in serum:0.2Serum level(mgml-1):0.03-0.4Complement activation:NoInteractions with cells:T cells via lectin like IgD receptorTransplacental transfer:NoIgA dimerisation and secretion IgA is the major isotype of antibody secreted at mucosal sufacesExists in serum as a monomer,but more usually as a J chain-linked dimer,that is formed in a similar manner to IgM pentamers.JCCSSSSCCSSSSCCs sIgA exists in two subclassesIgA1 is mostly found in serum and made by bone marrow B cells IgA2 is mostly found in mucosal secretions,colostrum and milk and is made by B cells located in the mucosae EpithelialcellJCCSSSSCCSSSSCCssSecretory IgA and transcytosisBJC CSSSSCCSSSSCCs sJCCSSSSCCSSSSCCssJCCSSSSCCSSSSCCsspIgR&IgA areinternalisedStalk of the pIgR is degraded to release IgA containing part of the pIgR-the secretory componentJCCSSSSCCSSSSCCssIgA and pIgR are transported to the apical surface in vesiclesB cells located in the submucosaproduce dimeric IgAPolymeric Ig receptors are expressed on the basolateral surface of epithelial cells to capture IgA produced in the mucosaIgA facts and figuresHeavy chains:a1 or a2-Alpha 1 or 2Half-life:IgA1 5-7 daysIgA2 4-6 daysSerum levels(mgml-1):IgA1 1.4-4.2IgA2 0.2-0.5%of Ig in serum:IgA1 11-14IgA2 1-4Complement activation:IgA1-by alternative and lectin pathwayIgA2-NoInteractions with cells:Epithelial cells by pIgRPhagocytes by IgA receptorTransplacental transfer:NoTo reduce vulnerability to microbial proteases the hinge region of IgA2 is truncated,and in IgA1 the hinge is heavily glycosylated.IgA is inefficient at causing inflammation and elicits protection by excluding,binding,cross-linking microorganisms and facilitating phagocytosisIgE facts and figuresIgE appears late in evolution in accordance with its role in protecting against parasite infectionsMost IgE is absorbed onto the high affinity IgE receptors of effector cellsIgE is also closely linked with allergic diseases Heavy chain:e-EpsilonHalf-life:1-5 daysSerum level(mgml-1):0.0001-0.0002%of Ig in serum:0.004Complement activation:NoInteractions with cells:Via high affinity IgE receptors expressed by mast cells,eosinophils,basophils and Langerhans cellsVia low affinity IgE receptor on B cells and monocytesTransplacental transfer:NoThe high affinity IgE receptor(Fce eRI)a chainb chaing2SSSSSSCe1Ce1Ce2Ce2Ce3Ce3Ce4Ce4Ce1Ce1Ce2Ce2Ce3Ce3Ce4Ce4The IgE-FceRI interaction is the highest affinity of any Fc receptor with an extremely low dissociation rate.Binding of IgE to FceRI increases the half life of IgECe3 of IgE interacts with the a chain of FceRI causing a conformational change.IgG facts and figuresHeavy chains:g 1 g 2 g3 g4-Gamma 1-4Half-life:IgG1 21-24 days IgG2 21-24 days IgG3 7-8 days IgG4 21-24 daysSerum level(mgml-1):IgG15-12IgG2 2-6IgG3 0.5-1IgG4 0.2-1%of Ig in serum:IgG145-53IgG2 11-15IgG3 3-6IgG4 1-4Complement activation:IgG1+IgG2+IgG3+IgG4 NoInteractions with cells:All subclasses via IgG receptors on macrophages and phagocytesTransplacental transfer:IgG1+IgG2+IgG3+IgG4+Carbohydrate is essential for complement activationSubtly different hinge regions between subclasses accounts for differing abilities to activate complement C1q binding motif is located on the Cg2 domainFcg g receptorsReceptorCell typeEffect of ligationFcg gRIMacrophages Neutrophils,Eosinophils,Dendritic cells Uptake,Respiratory burstFcg gRIIAMacrophages Neutrophils,Eosinophils,PlateletsLangerhans cells Uptake,Granule releaseFcg gRIIB1 B cells,Mast CellsNo Uptake,Inhibition of stimulationFcg gRIIB2 Macrophages Neutrophils,Eosinophils Uptake,Inhibition of stimulationFcg gRIIINK cells,Eosinophils,Macrophages,NeutrophilsMast cellsInduction of killing(NK cells)High affinity Fcg receptors from the Ig superfamily:The neonatal Fcg g receptorThe FcgRn is structurally related to MHC class IIn cows FcgRn binds maternal IgG in the colostrum at pH 6.5 in the gut.The IgG receptor complex is trancytosed across the gut epithelium and the IgG is released into the foetal blood by the sharp change in pH to 7.4Some evidence that this may also happen in the human placenta,however the mechanism is not straightforward.Human FcgRnHuman MHCClass I