All complexes of T cell receptors (TCRs) bound to peptide-major histocompatibility complex (pMHC) molecules assume a stereotyped binding 'polarity', despite wide variations in TCR-pMHC docking angles. However, existing TCR-pMHC crystal structures have failed to show broadly conserved pairwise interaction motifs. Here we determined the crystal structures of two TCRs encoded by the variable beta-chain 8.2 (V(beta)8.2), each bound to the MHC class II molecule I-A(u), and did energetic mapping of V(alpha) and V(beta) contacts with I-A(u). Together with two previously solved structures of V(beta)8.2-containing TCR-MHC complexes, we found four TCR-I-A complexes with structurally superimposable interactions between the V(beta) loops and the I-A alpha-helix. This examination of a narrow 'slice' of the TCR-MHC repertoire demonstrates what is probably one of many germline-derived TCR-MHC interaction 'codons'.

HLA-B * 4402 and B * 4403 are naturally occurring MHC class I alleles that are both found at a high frequency in all human populations, and yet they only differ by one residue on the ␣ 2 helix (B * 4402 Asp156 → B * 4403 Leu156). CTLs discriminate between HLA-B * 4402 and B * 4403, and these allotypes stimulate strong mutual allogeneic responses reflecting their known barrier to hemopoeitic stem cell transplantation. Although HLA-B * 4402 and B * 4403 share Ͼ 95% of their peptide repertoire, B * 4403 presents more unique peptides than B * 4402, consistent with the stronger T cell alloreactivity observed toward B * 4403 compared with B * 4402. Crystal structures of B * 4402 and B * 4403 show how the polymorphism at position 156 is completely buried and yet alters both the peptide and the heavy chain conformation, relaxing ligand selection by B * 4403 compared with B * 4402. Thus, the polymorphism between HLA-B * 4402 and B * 4403 modifies both peptide repertoire and T cell recognition, and is reflected in the paradoxically powerful alloreactivity that occurs across this "minimal" mismatch. The findings suggest that these closely related class I genes are maintained in diverse human populations through their differential impact on the selection of peptide ligands and the T cell repertoire.Key words: class I histocompatibility molecules • antigen presentation • crystallography • X-ray diffraction • graft rejection • polymorphism protective immunity against microbes (1-3). HLA alleles can differ from each other by only a single amino acid ("micropolymorphism") or by Ͼ 30 amino acids (4). It has been suggested that there are nine major HLA class I "supertypes," or clusters of alleles, that each possess a unique broad specificity for common anchor motifs in antigenic peptides (5). Alleles from each of these supertypic families are distributed in virtually all human populations and account

T helper type 17 (TH-17) cells, together with their effector cytokines including interleukin 17 (IL-17) family members, are emerging as key mediators of chronic inflammatory and autoimmune disorders. Here we present the crystal structure of a 1:2 complex of IL-17RA bound to IL-17F. The manner of complex formation is unique for cytokines, and involves two fibronectin-type domains of IL-17RA engaging IL-17 within a groove between the IL-17 homodimer interface in a knob-and-hole fashion. The first receptor-binding event to the IL-17 cytokines modulates the affinity and specificity of the second receptor-binding event, thereby promoting heterodimeric versus homodimeric complex formation. IL-17RA utilizes a common recognition strategy to bind to several IL-17 family members, allowing it to potentially act as a shared receptor within multiple different signaling complexes.

HLA class I polymorphism creates diversity in epitope specificity and T cell repertoire. We show that HLA polymorphism also controls the choice of Ag presentation pathway. A single amino acid polymorphism that distinguishes HLA-B*4402 (Asp116) from B*4405 (Tyr116) permits B*4405 to constitutively acquire peptides without any detectable incorporation into the transporter associated with Ag presentation (TAP)-associated peptide loading complex even under conditions of extreme peptide starvation. This mode of peptide capture is less susceptible to viral interference than the conventional loading pathway used by HLA-B*4402 that involves assembly of class I molecules within the peptide loading complex. Thus, B*4402 and B*4405 are at opposite extremes of a natural spectrum in HLA class I dependence on the PLC for Ag presentation. These findings unveil a new layer of MHC polymorphism that affects the generic pathway of Ag loading, revealing an unsuspected evolutionary trade-off in selection for optimal HLA class I loading versus effective pathogen evasion.

A human anti-IL-2 antibody that potentiates regulatory T cells by a structure-based mechanism . Previous studies showed that IL-2 is highly flexible 8,9 and exists in different conformations that favor either the high-affinity trimeric IL-2R or intermediate-affinity dimeric IL-2R, resulting in the activation of different immune cells 9 . This plasticity has complicated the use of the approved drug Proleukin at high doses to treat metastatic melanoma and renal cell carcinoma 10 , due to the role of IL-2 as an essential growth factor for T regs [11][12][13] . Moreover, adverse effects of high-dose IL-2 therapy have greatly limited its use 14,15 . Several studies have shown that low-dose IL-2 therapy preferentially activates T regs due to the constitutive high expression of IL-2Rα 16 and other cell-intrinsic factors that increase signal transduction sensitivity 17 . Treatment of mice and humans with low doses of IL-2 has been shown to ameliorate autoimmune diseases and graft-versus-host disease (GvHD) as well as delaying organ allograft rejection [18][19][20][21][22] . However, IL-2 therapy has some limitations, including difficulty in predicting the efficacious dose, off-target effects on different cell populations and a short in vivo half-life 23,24 . Thus, attempts have been made to engineer or modify the IL-2 structure to improve its therapeutic potential by modulating its ability to selectively target either T effs or T regs [25][26][27][28][29][30][31] . Selective antibodies against IL-2 can alter its conformation by binding a number of potential epitopes, thereby modifying the binding interaction of IL-2 to any of the IL-2R subunits and resulting in selective expansion of T regs or T eff cell subsets 32,33 . For example, it has been demonstrated that a rat anti-mouse IL-2 monoclonal antibody (JES6-1) can be administered in complex with wild-type mouse IL-2 and used to preferentially enhance T reg populations 26 . Binding of JES6-1 to IL-2 alters its conformation to lower the affinity of mIL-2 for CD25, such that CD25 high T regs compete favorably for IL-2 binding and expansion against T effs 33 . The therapeutic potential of IL-2 to selectively activate the tolerogenic immune response, combined with the imperative to develop a human T reg -selective IL-2 compound, led us to develop a mechanism-based screening strategy to identify human antibodies against human IL-2 that exhibit an in vivo T reg potentiation profile when complexed with hIL-2. This class of monoclonal antibody, exemplified by F5111.2, blocked IL-2Rβ binding and reduced IL-2Rα binding to IL-2, and, when administered in complex with hIL-2, preferentially promoted T reg expansion and was effective in models of autoimmune disease including type 1 diabetes and experimental autoimmune encephalomyelitis (EAE) in addition to GvHD. ResultsSelective IL-2 stimulation in T regs . To directly compare T reg and T eff sensitivity to IL-2, the pSTAT5 signaling response of T regs was analyzed in a mixed population of peripheral blood mononuclear cells (PBMCs).

The CD3␥ heterodimer is essential for expression and function of the T cell receptor. The crystal structure of the human CD3␥ heterodimer is described to 2.1-Å resolution complexed with OKT3, a therapeutic mAb that not only activates and tolerizes mature T cells but also induces regulatory T cells. The mode of CD3␥ dimerization provides a general structural basis for CD3 assembly and maps candidate T cell antigen receptor docking sites, including a duplicated linear region rich in acidic residues that is unique to human CD3. OKT3 binds to an atypically small area of CD3 and has a low affinity for the isolated CD3␥ heterodimer. The structure of the OKT3͞CD3␥ complex has implications for T cell signaling and therapeutic design. T cell recognition is mediated by clonotypically distributed ␣␤ and ␥␦ T cell receptors (TcR) that interact with the peptideloaded molecules of the peptide MHC (pMHC) (1). The antigenspecific chains of the TcR do not possess signaling domains but instead are coupled to the conserved multisubunit signaling apparatus CD3 (2-4). The mechanism by which TcR ligation is directly communicated to the signaling apparatus remains a fundamental question in T cell biology (3, 5). It seems clear that sustained T cell responses involve coreceptor engagement, TcR oligomerization, and a higher order arrangement of TcR-pMHC complexes in the immunological synapse (6-9). However very early TcR signaling occurs in the absence of these events and may involve a ligandinduced conformational change in CD3 (3, 5, 10, 11). The , ␥, ␦ and subunits of the signaling complex associate with each other to form a CD3␥ heterodimer, a CD3␦ heterodimer, and a CD3 homodimer (2). Transfection studies (12), gene knockouts (13), and natural mutations (14) have revealed that the CD3 molecules are important for the proper cell surface expression of the ␣␤ TcR and normal T cell development. The solution structure of the ectodomain fragments of the mouse CD3␥ heterodimer showed that the ␥ subunits are both C2-set Ig domains that interact with each other to form an unusual side-to-side dimer configuration (15). Although the cysteine-rich stalk appears to play an important role in driving CD3 dimerization (15, 16), interaction by means of the extracellular domains of CD3 and CD3␥ is sufficient for assembly of these proteins with TcR␤ (17, 18). Although still controversial, the dominant stoichiometry of the TcR most likely comprises one ␣␤ TcR, one CD3␥ heterodimer, one CD3␦ heterodimer and one CD3 homodimer (2).A number of therapeutic strategies modulate T cell immunity by targeting TcR signaling, particularly the anti-human CD3 mAbs that are widely used clinically in immunosuppressive regimes. The CD3-specific mouse mAb OKT3 was the first mAb licensed for use in humans (19) and is widely used clinically as an immunosuppressive agent in transplantation (20-23), type 1 diabetes (21, 24), and psoriasis (25). Moreover, nonmitogenic anti-CD3 mAbs can induce partial T cell signaling and clonal anergy (26). Anti-CD3 treatment is also reporte...

The elusive etiology of germline bias of the T cell receptor (TCR) for major histocompatibility complex (MHC) has been clarified by recent ‘proof-of-concept’ structural results demonstrating the conservation of specific TCR-MHC interfacial contacts in complexes bearing common variable segments and MHC allotypes. We suggest that each TCR variable-region gene product engages each type of MHC through a ‘menu’ of structurally coded recognition motifs that have arisen through coevolution. The requirement for MHC-restricted T cell recognition during thymic selection and peripheral surveillance has necessitated the existence of such a coded recognition system. Given these findings, a reconsideration of the TCR–peptide-MHC structural database shows that not only have the answers been there all along but also they were predictable by the first principles of physical chemistry.

The energetic bases of T cell recognition are unclear. Here, we studied the 'energetic landscape' of peptide-major histocompatibility complex (pMHC) recognition by an immunodominant alphabeta T cell receptor (TCR). We quantified and evaluated the effect of natural and systematic substitutions in the complementarity-determining region (CDR) loops on ligand binding in the context of the structural detail of each component of the immunodominant TCR-pMHC complex. The CDR1 and CDR2 loops contributed minimal energy through direct recognition of the antigen and instead had a chief function in stabilizing the ligated CDR3 loops. The underlying energetic basis for recognition lay in the CDR3 loops. Therefore the energetic burden of the CDR loops in the TCR-pMHC interaction is variable among TCRs, reflecting the inherent adaptability of the TCR in ligating different ligands.