8536d_cho7 161-184 8/16/02 12: 09 PM Page 167 mac100 mac 100: 1?/8_tm: 8536d: Goldsby et al./Immunology Se Major Histocompatibility Complex CHAPTER 7 Peptide-binding cleft a1 domain a 2 domain Sheets a2 domain g a3 domain FIGURE7-6Representations of the three-dimensional structure of munoglobulin- fold structure of the a3 domain and B2-microglobulin the external domains of a human class I MHC molecule based on x. (b) The al and a2 domains as viewed from the top, showing the ray crystallographic analysis. (a) Side view in which the B strands are peptide-binding cleft consisting of a base of antiparallel p strands depicted as thick arrows and the a helices as spiral ribbons. Disulfide and sides of a helices. this cleft in class I molecules can accommo- bonds are shown as two interconnected spheres. The al and a2 do- date peptides containing 8-10 residues ding cleft. Note the im- domain,B2-microglobulin, and the constant-region domains which is not surprising given the considerable sequence sim- in immunoglobulins. The enzyme papain cleaves the a chain ilarity with the immunoglobulin constant regions, class I just 13 residues proximal to its transmembrane domain, re- MHC molecules and B2-microglobulin are classified as leasing the extracellular portion of the molecule, consisting of members of the immunoglobulin superfamily( see Figure al,a2, a3, and B2-microglobulin. Purification and crystal- 4-20). The a3 domain appears to be highly conserved among lization of the extracellular portion revealed two pairs of in- class I MHC molecules and contains a sequence that interacts teracting domains: a membrane-distal pair made up of the al with the CD8 membrane molecule present on Tc cells and a2 domains and a membrane-proximal pair composed of B2-Microglobulin interacts extensively with the a3 do- the a3 domain and B2-microglobulin( Figure 7-6a) main and also interacts with amino acids of the al and a2 The al and o2 domains interact to form a platform of domains. The interaction of B2-microglobulin and a peptide eight antiparallel B strands spanned by two long a-helical re- with a class I a chain is essential for the class I molecule to gions. The structure forms a deep groove, or cleft, approxi- reach its fully folded conformation As described in detail in mately 25 A X 10 A X 11 A, with the long a helices as sides Chapter 8, assembly of class I molecules is believed to occur nd the B strands of the B sheet as the bottom( Figure 7-6b). by the initial interaction of B2-microglobulin with the fold This peptide-binding cleft is located on the top surface of the ing class I a chain. This metastable"empty dimer is then sta- class I MHC molecule, and it is large enough to bind a peptide bilized by the binding of an appropriate peptide to form the of 8-10 amino acids. The great surprise in the x-ray crystallo- native trimeric class I structure consisting of the class I a graphic analysis of class I molecules was the finding of small chain, B2-microglobulin, and a peptide. This complete mole peptides in the cleft that had cocrystallized with the protein. cular complex is ultimately transported to the cell surface These peptides are, in fact, processed antigen and self-pep In the absence of B2-microglobulin, the class I MHC tides bound to the al and a2 domains in this deep groove. chain is not expressed on the cell membrane. This is illus The a3 domain and B2-microglobulin are organized into trated by Daudi tumor cells, which are unable to synthesize two p pleated sheets each formed by antiparallel p strands of B2-microglobulin. These tumor cells produce class I MHCa amino acids. As described in Chapter 4, this structure, known chains, but do not express them on the membrane. However, as the immunoglobulin fold, is characteristic of im- if Daudi cells are transfected with a functional gene encoding munoglobulin domains. Because of this structural similarity, B2-microglobulin, class I molecules appear on the membranedomain, 2-microglobulin, and the constant-region domains in immunoglobulins. The enzyme papain cleaves the chain just 13 residues proximal to its transmembrane domain, re￾leasing the extracellular portion of the molecule, consisting of 1, 2, 3, and 2-microglobulin. Purification and crystal￾lization of the extracellular portion revealed two pairs of in￾teracting domains: a membrane-distal pair made up of the 1 and 2 domains and a membrane-proximal pair composed of the 3 domain and 2-microglobulin (Figure 7-6a). The 1 and 2 domains interact to form a platform of eight antiparallel  strands spanned by two long -helical re￾gions. The structure forms a deep groove, or cleft, approxi￾mately 25 Å  10 Å  11 Å, with the long helices as sides and the  strands of the  sheet as the bottom (Figure 7-6b). This peptide-binding cleft is located on the top surface of the class I MHC molecule, and it is large enough to bind a peptide of 8–10 amino acids. The great surprise in the x-ray crystallo￾graphic analysis of class I molecules was the finding of small peptides in the cleft that had cocrystallized with the protein. These peptides are, in fact, processed antigen and self-pep￾tides bound to the 1 and 2 domains in this deep groove. The 3 domain and 2-microglobulin are organized into two  pleated sheets each formed by antiparallel  strands of amino acids. As described in Chapter 4, this structure, known as the immunoglobulin fold, is characteristic of im￾munoglobulin domains. Because of this structural similarity, which is not surprising given the considerable sequence sim￾ilarity with the immunoglobulin constant regions, class I MHC molecules and 2-microglobulin are classified as members of the immunoglobulin superfamily (see Figure 4-20). The 3 domain appears to be highly conserved among class I MHC molecules and contains a sequence that interacts with the CD8 membrane molecule present on TC cells. 2-Microglobulin interacts extensively with the 3 do￾main and also interacts with amino acids of the 1 and 2 domains. The interaction of 2-microglobulin and a peptide with a class I chain is essential for the class I molecule to reach its fully folded conformation. As described in detail in Chapter 8, assembly of class I molecules is believed to occur by the initial interaction of 2-microglobulin with the fold￾ing class I chain. This metastable “empty” dimer is then sta￾bilized by the binding of an appropriate peptide to form the native trimeric class I structure consisting of the class I chain, 2-microglobulin, and a peptide. This complete mole￾cular complex is ultimately transported to the cell surface. In the absence of 2-microglobulin, the class I MHC chain is not expressed on the cell membrane. This is illus￾trated by Daudi tumor cells, which are unable to synthesize 2-microglobulin. These tumor cells produce class I MHC chains, but do not express them on the membrane. However, if Daudi cells are transfected with a functional gene encoding 2-microglobulin, class I molecules appear on the membrane. Major Histocompatibility Complex CHAPTER 7 167 (b) α1 domain α2 domain α3 domain α2 domain α1 domain β2-microglobulin α helix β sheets (a) Peptide-binding cleft FIGURE 7-6 Representations of the three-dimensional structure of the external domains of a human class I MHC molecule based on x￾ray crystallographic analysis. (a) Side view in which the  strands are depicted as thick arrows and the helices as spiral ribbons. Disulfide bonds are shown as two interconnected spheres. The 1 and 2 do￾mains interact to form the peptide-binding cleft. Note the im￾munoglobulin-fold structure of the 3 domain and 2-microglobulin. (b) The 1 and 2 domains as viewed from the top, showing the peptide-binding cleft consisting of a base of antiparallel  strands and sides of helices. This cleft in class I molecules can accommo￾date peptides containing 8–10 residues. 8536d_ch07_161-184 8/16/02 12:09 PM Page 167 mac100 mac 100: 1268_tm:8536d:Goldsby et al. / Immunology 5e-: