Antibodies and Antigens Antibodies and Antigens Your body’ ’s natural defense s natural defense Your body system system Antibodies Antibodies The formal chemical name for antibodies The formal chemical name for antibodies is immunoglobulins immunoglobulins. . is Immunoglobulins are protein molecules are protein molecules Immunoglobulins which defend your body from foreign which defend your body from foreign toxins known as toxins known as antigens antigens . . Immunoglobulin G, IgG, is the major Immunoglobulin G, IgG, is the major antibody in serum (blood) and will be what antibody in serum (blood) and will be what we use for example purposes. we use for example purposes. 1
Immunoglobulin Structure Immunoglobulin Structure http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stryer.figgrp.4700 IgG, is comprised of 3 polypeptide chains IgG, is comprised of 3 polypeptide chains � 4 4 “ “light light” ” chains, L chains, L – – 25 25- -kd chains (shown in yellow) kd chains (shown in yellow) � � 2 2 “ “heavy heavy” ” chain, H chain, H – – 50 50- -kd chain (shown in blue) kd chain (shown in blue) � A disulfide bond links L to H. A disulfide bond links L to H. S S H chains link to each other by 1or more disulfide bonds. H chains link to each other by 1or more disulfide bonds. Immunoglobulin Cleavage Immunoglobulin Cleavage IgG is cleaved by an enzyme IgG is cleaved by an enzyme called papain papain into 3 fragments, into 3 fragments, called F. F. F ab = antigen binding fragment F ab = antigen binding fragment � � F c F c = constant fragment (does = constant fragment (does � � not bind to antigens) not bind to antigens) The disulfide bonds connecting The disulfide bonds connecting F F ab ab and F and F c c are flexible and are flexible and called called “ “hinges hinges” ” http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stryer.figgrp.4701 This flexibility allows for one This flexibility allows for one � � antibody to bind to antigens antibody to bind to antigens with multiple bonding sites. with multiple bonding sites. http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stryer.figgrp.4703 2
Antigen Cross- Antigen Cross -linking linking Since each IgG Since each IgG contains two F ab contains two F ab chains it can cross- - chains it can cross link multiple antigens link multiple antigens http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stryer.figgrp.4702 Other Immunoglobulins Other Immunoglobulins IgA – IgA – external secretions: saliva, tears, mucus, etc. external secretions: saliva, tears, mucus, etc. IgM – – appears first in serum upon introduction of antigen, 10 binding appears first in serum upon introduction of antigen, 10 binding sites sites IgM allows for excellent binding to multiple receptor antigens allows for excellent binding to multiple receptor antigens IgD IgD – – role not known role not known IgE – – protection from parasites, can also cause allergic reactions protection from parasites, can also cause allergic reactions IgE http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stryer.figgrp.4704 3
Variable Regions – – Mechanism of Mechanism of Variable Regions Antibody Action Antibody Action The end terminus of the The end terminus of the F ab chains are HIGHLY F ab chains are HIGHLY variable. This is why our variable. This is why our bodies are able to form bodies are able to form antibodies to nearly all antibodies to nearly all antigens. antigens. � V V L = variable light L = variable light � � V V H = variable heavy H = variable heavy � � C C L = constant light L = constant light � � C C H = constant heavy H = constant heavy � Why can they vary so Why can they vary so widely? http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stryer.figgrp.4707 widely? V L and V H Chemistry – – the the REAL REAL V L and V H Chemistry BASICS! BASICS! V V L L and V and V H H are complex are complex proteins which adopt a proteins which adopt a common structure called common structure called the immunoglobulin fold. immunoglobulin fold. the The immunoglobulin The immunoglobulin fold fold is constructed from is constructed from β - - sheets. The beta two β two sheets. The beta http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stryer.figgrp.4709 sheets contain anti sheets contain anti- - Key aspect: Key aspect: parallel strands. These parallel strands. These Three loops at the end are Three loops at the end are � � “ hypervariable, “ hypervariable,” ” meaning that their meaning that their strands surround a strands surround a amino acid sequences can be varied amino acid sequences can be varied hydrophobic core. The hydrophobic core. The in many, many ways. in many, many ways. two sheets are connected two sheets are connected These loops are the variable regions These loops are the variable regions � � we same in the previous slide. we same in the previous slide. via disulfide bond. via disulfide bond. This region is also known as the This region is also known as the � � complementarity determining region complementarity determining region or or CDR. CDR. 4
Big Picture Big Picture So if V H can vary immensely and V L can very So if V H can vary immensely and V L can very immensely then: immensely then: i,j ∞ Σ (V L ) i (V H ) j + = i,j=1 Ok, so more like a really big number! How’ ’d he do that? d he do that? How So then you say “ “How can our body make How can our body make So then you say all these different antibodies?” ” all these different antibodies? I say : “ “That is what B That is what B- -cells are for! cells are for!” ” I say : 5
B B- -cells and T cells and T- -cells cells Both develop in the bone Both develop in the bone marrow marrow B B- -cells mature in the cells mature in the B one marrow as well one marrow as well B whereas, T- -cells mature cells mature whereas, T in the in the T T hymus. hymus. B- -cells produce cells produce B antibodies with the help antibodies with the help of T- -cells. cells. of T Some T- Some T -cells can act as cells can act as antibodies themselves antibodies themselves (i.e., against viruses but (i.e., against viruses but we won’ ’t get into that) t get into that) we won http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/B/B_and_Tcells.html Why do we care and how the heck Why do we care and how the heck does this relate to Chemistry? does this relate to Chemistry? #1 Everything relates to CHEMISTRY CHEMISTRY !!! !!! #1 Everything relates to #2 #2 – – You should care because this is how You should care because this is how your body protects itself. your body protects itself. #3 – – One of the many ways this relates to One of the many ways this relates to #3 Chemistry is the relationship between Chemistry is the relationship between antigens and antibodies both kinetically kinetically antigens and antibodies both and in terms of equilibrium and in terms of equilibrium . . 6
Antibody/Antigen Affinity Antibody/Antigen Affinity A very important aspect of the relationship A very important aspect of the relationship between antibodies and antigens is their between antibodies and antigens is their affinities for each other. Some affinities for each other. Some antibody/antigen pairs have a relatively antibody/antigen pairs have a relatively high affinity for each other and some have high affinity for each other and some have a somewhat lower affinity for each other. a somewhat lower affinity for each other. Equilibrium Equilibrium Think back to when we studied Think back to when we studied equilibrium. equilibrium. We showed that when species are We showed that when species are associated with each other we can associated with each other we can measure the degree of association with an measure the degree of association with an ???? ???? Equilibrium Constant, K Equilibrium Constant, K 7
Example Example We’ ’ll call antigens G and antibodies A. ll call antigens G and antibodies A. We K K d d stands for Equilibrium of dissociation stands for Equilibrium of dissociation ⇔ + GA G A [ ][ ] G A d = K [ ] GA Try these problems! Try these problems! Suppose that the dissociation constant of an Suppose that the dissociation constant of an 7 M at 25 ° C. M at 25 ° F ab -hapten complex is 3 x 10 hapten complex is 3 x 10 - -7 C. F ab - A) Immunologist often speak of affinity (K A) Immunologist often speak of affinity (K a a ), the ), the reciprocal of the dissociation constant, in comparing reciprocal of the dissociation constant, in comparing antibodies. What is the affinity of this F ab antibodies. What is the affinity of this F ab ? ? B) What is the standard free energy of binding? What is the standard free energy of binding? B) C) The rate constant of release of hapten from the The rate constant of release of hapten from the C) complex is 120 s - -1 1 . What is the rate constant of . What is the rate constant of complex is 120 s association? association? http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stryer.section.4770 8
A. A. 1 = K a K d 1 = = × − 6 1 3 . 3 10 K M − × a 7 3 10 M B. B. Δ = − o ln G RT K f for binding this would be K a ⋅ J mol ( ) Δ = − × 6 − 1 o 8 . 3145 298 ln( 3 . 3 10 ) G K M f K Δ = − o 37214 G J f Δ = − 37 o G kJ f 9
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