IEDB-3D: structural data within the immune epitope database

From the initial observations by Zinkernagel and Doherty over 40 years ago of the need for T cells to recognize "altered self," our understanding of TCR recognition of peptide+ MHC has made significant advances. However, alongside a more detailed understanding of the interaction comes additional questions around precisely why T cells must limit themselves to MHC, when a greater range of ligand binding is demonstrably possible, and mechanistically, how MHC restriction achieves the necessary T cell survival and activation signals. The answer may well lie in the study of noncanonical or poorly signaling TCRs to understand the absolute requirements for effective TCR-pMHC recognition, continued advances in structural biology providing resolution of multimolecular complexes, and cryo-EM providing information on the dynamics of molecular localization and organization before and after TCR ligation of pMHC.

BackgroundStructural information about epitopes, particularly the three-dimensional (3D) structures of antigens in complex with immune receptors, presents a valuable source of data for immunology. This information is available in the Protein Data Bank (PDB) and provided in curated form by the Immune Epitope Database and Analysis Resource (IEDB). With continued growth in these data and the importance in understanding molecular level interactions of immunological interest there is a need for new specialized molecular visualization and analysis tools.ResultsThe EpitopeViewer is a platform-independent Java application for the visualization of the three-dimensional structure and sequence of epitopes and analyses of their interactions with antigen-specific receptors of the immune system (antibodies, T cell receptors and MHC molecules). The viewer renders both 3D views and two-dimensional plots of intermolecular interactions between the antigen and receptor(s) by reading curated data from the IEDB and/or calculated on-the-fly from atom coordinates from the PDB. The 3D views and associated interactions can be saved for future use and publication. The EpitopeViewer can be accessed from the IEDB Web site through the quick link 'Browse Records by 3D Structure.'ConclusionThe EpitopeViewer is designed and been tested for use by immunologists with little or no training in molecular graphics. The EpitopeViewer can be launched from most popular Web browsers without user intervention. A Java Runtime Environment (RJE) 1.4.2 or higher is required.

Peptides presented by the major histocompatibility complex (MHC) are derived from the degradation of cellular proteins. Thus, the repertoire of these peptides (the MHC peptidome) should correlate better with the cellular protein degradation scheme (the degradome) than with the cellular proteome. To test the validity of this statement and to determine whether the majority of MHC peptides are derived from short lived proteins, from defective ribosome products, or from regular long lived cellular proteins we analyzed in parallel the turnover kinetics of both MHC peptides and cellular proteins in the same cancer cells. The analysis was performed by pulse-chase experiments based on stable isotope labeling in tissue culture followed by capillary chromatography and tandem mass spectrometry. Indeed only a limited correlation was observed between the proteome and the MHC peptidome observed in the same cells. Moreover a detailed analysis of the turnover kinetics of the MHC peptides helped to assign their origin to normal, to short lived or long lived proteins, or to the defective ribosome products. Furthermore the analysis of the MHC peptides turnover kinetics helped to direct attention to abnormalities in the degradation schemes of their source proteins. These observations can be extended to search for cancer-related abnormalities in protein degradation, including those that lead to loss of tumor suppressors and cell cycle regulatory proteins.

Major Histocompatibility Complex (MHC) molecules play a crucial role in inducing an adaptive immune response. T-cell epitopes require compatible MHC molecules to form MHC-peptide Complexes (pMHC) that activate the T-cell Receptors (TCR) of T-lymphocyte clones. MHCs are polymorphic molecules with wide varieties of gene alleles. There are two classes of MHC molecules, class I and II. Both classes have three classical loci HLA-A, -B, and –C are present in class I and HLA-DP, -DQ, and -DR in class II. To induce a compatible T-lymphocyte clone, the T-cell epitope requires the association of the compatible MHC molecule to form pMHC. Each MHC variant possesses a different groove that is capable of binding a different range of antigenic epitopes. Without the compatible MHC molecule, a T cell clone cannot be activated by a particular viral epitope. With the aim of preventing viral transmission, the efficiency of a viral vaccine is related to the existence of specific MHC alleles in the individual. This article proposes the roles of the MHC molecule to prevent viral infection. In addition, the association of the viral receptor molecule with the viral infection will also be discussed.

The U21 gene product from human herpesvirus 7 binds to and redirects class I major histocompatibility complex (MHC) molecules to a lysosomal compartment. The molecular mechanism by which U21 reroutes class I MHC molecules to lysosomes is not known. Here, we have reconstituted the interaction between purified soluble U21 and class I MHC molecules, suggesting that U21 does not require additional cellular proteins to interact with class I MHC molecules. Our results demonstrate that U21, itself predicted to contain an MHC class I-like protein fold, interacts tightly with class I MHC molecules as a tetramer, in a 4:2 stoichiometry. These observations have helped to elucidate a refined model describing the mechanism by which U21 escorts class I MHC molecules to the lysosomal compartment. In this report, we show that the human herpesvirus 7 (HHV-7) immunoevasin U21, itself a class I MHC-like protein, binds with high affinity to class I MHC molecules as a tetramer and escorts them to lysosomes, where they are degraded. While many class I MHC-like molecules have been described in detail, this unusual viral class I-like protein functions as a tetramer, associating with class I MHC molecules in a 4:2 ratio, illuminating a functional significance of homooligomerization of a class I MHC-like protein.

The adaptive immune system in vertebrates has evolved to recognize an immense number of pathogens. B and T cells play important role in the antigen recognition by expressing a substantially diverse repertoire of antigen-specific receptors. The recent advances in the high-throughput immune repertoire sequencing has led to a tremendous increase in the antibody and T-cell receptor information. Identification of specific epitopes recognized by the receptors is vital in analyzing the repertoire sequencing data. Here, we present a tool that examines data from an immune receptor repertoire experiment, and identifies receptors (or highly related variants) that are present in the Immune Epitope Database (IEDB). The IEDB is a free resource which compiles experiments characterizing immune epitopes. IEDB also captures the epitope-specific immune receptor sequences and groups them based on their epitope specificity. IEDB currently has information on over 26,000 receptor groups. Users can provide receptor repertoire sequence data in multiple formats, and specify different CDR sequence identity cutoffs to restrict or broaden the receptor search. We expect that this tool will provide new insights into B and T cell responses captured in the receptor repertoire experiments by identifying specific epitopes, and facilitate development of new strategies for monitoring and treating of infectious, allergic and autoimmune diseases.

Herpesviruses have evolved numerous strategies to evade detection by the immune system. Notably, most of the herpesviruses interfere with viral antigen presentation to cytotoxic T lymphocytes (CTLs) by removing class I major histocompatibility complex (MHC) molecules from the infected cell surface. Clearly, since the herpesviruses have evolved an extensive array of mechanisms to remove class I MHC molecules from the cell surface, this strategy serves them well. However, class I MHC molecules often serve as inhibitory ligands for NK cells, so viral downregulation of all class I MHC molecules should leave the infected cell open to NK cell attack. Some viruses solve this problem by selectively downregulating certain class I MHC products, leaving other class I products at the cell surface to serve as inhibitory NK cell ligands. Here, we show that human herpesvirus 7 (HHV-7) U21 binds to and downregulates all of the human class I MHC gene products, as well as the murine class I molecule H-2K(b). HHV-7-infected cells must therefore possess other means of escaping NK cell detection.

An antigen will be able to stimulate an immune response if the antigen has been presented and recognized by T-cells. Major Histocompatibility Complex (MHC) molecules are molecules that will recognize and present the peptides of the antigen. In humans, two classes of MHC are known to carry out this role, namely Class-I MHC and Class-II MHC, while Class-III MHC has a function that is not directly related to peptide binding. The genes that code for MHC molecules are so polymorphic that each individual is certain to have a different allele (heterozygous). In Class-I MHC, only the α-chain is polymorphic while in Class-II MHC both α and β-chains are polymorphic. MHC polymorphism will affect (1) the ability to create immune responses including antibody production, (2) resistance or susceptibility to infectious diseases, and (3) resistance or susceptibility to autoimmune and allergic diseases. That is why MHC is so polymorphic. The immune system has to fight many different pathogens. By having many different MHC molecules, one can present widely different antigens and can spur an effective immune response.

Neoantigens are tumor-derived peptides and are biomarkers that can predict prognosis related to immune checkpoint inhibition by estimating their binding to major histocompatibility complex (MHC) proteins. Although deep neural networks have been primarily used for these prediction models, it is difficult to interpret the models reported thus far as accurately representing the interactions between biomolecules. In this study, we propose the GraphMHC model, which utilizes a graph neural network model applied to molecular structure to simulate the binding between MHC proteins and peptide sequences. Amino acid sequences sourced from the immune epitope database (IEDB) undergo conversion into molecular structures. Subsequently, atomic intrinsic informations and inter-atomic connections are extracted and structured as a graph representation. Stacked graph attention and convolution layers comprise the GraphMHC network which classifies bindings. The prediction results from the test set using the GraphMHC model showed a high performance with an area under the receiver operating characteristic curve of 92.2% (91.9-92.5%), surpassing a baseline model. Moreover, by applying the GraphMHC model to melanoma patient data from The Cancer Genome Atlas project, we found a borderline difference (0.061) in overall survival and a significant difference in stromal score between the high and low neoantigen load groups. This distinction was not present in the baseline model. This study presents the first feature-intrinsic method based on biochemical molecular structure for modeling the binding between MHC protein sequences and neoantigen candidate peptide sequences. This model can provide highly accurate responsibility information that can predict the prognosis of immune checkpoint inhibitors to cancer patients who want to apply it.

The exact role of major histocompatibility complex (MHC) molecules in the peripheral survival of naive T cells is controversial, as some studies have suggested that they are critically required whereas others have suggested that they are not. Here we controlled for some of the features that differed among the earlier studies, and analyzed both the survival and expansion of naive CD4+ T cells transferred into MHC syngeneic, allogeneic, or MHC negative environments. We found that naive T cells transferred into MHC negative or allogeneic environments often fail to survive because of rejection and/or competition by natural killer (NK) cells, rather than failure to recognize a particular MHC allele. In the absence of NK cells, naive CD4+ T cells survived equally well regardless of the MHC type of the host. There was, however, an MHC requirement for extensive space-induced “homeostatic” expansion. Although the first few divisions occurred in the absence of MHC molecules, the cells did not continue to divide or transit to a CD44hi phenotype. Surprisingly, this MHC requirement could be satisfied by alleles other than the restricting haplotype. Therefore, space-induced expansion and survival are two different phenomena displaying different MHC requirements. Memory CD4+ T cells, whose survival and expansion showed no requirements for MHC molecules at all, dampened the space-induced expansion of naive cells, showing that the two populations are not independent in their requirements for peripheral niches.

After viral infection, CD8+ T-cells can rapidly detect their intracellular presence by recognizing peptide fragments presented by MHC class I molecules on the cell surface that are derived from degraded viral protein. This appears to be in conflict with the observation that viral proteins have relatively long half-lives. To explain this, the DRiPs hypothesis (Yewdell and Nicchitta, 2006) was proposed, which states that these epitopes come from Defective Ribosomal Products (DRiPs), rather than degradation of mature protein products. If epitopes do come from defective ribosomal products, this could lead to a positional bias of epitopes in their corresponding antigens. Specifically, if a major source of defective ribosomal products is pre-mature stop in translation, an overrepresentation of CD8+ T-cell epitopes in the N-terminal portion of antigens would be expected. In contrast, if downstream initiation is a major source of defective ribosomal products, C-terminal portions of antigens would be a dominant source of epitopes. In this study, we systematically assess positional bias of epitopes in viral antigens, taking advantage of the large set of immune epitope data available in the Immune Epitope Database. We demonstrate that there is a small degree of positional skewing detectable in the epitope distribution. Epitopes from both ends of antigens tend to be slightly underrepresented, which suggests that both pre-mature stops in translation and downstream initiation play an equal but minor role in shaping the overall viral epitope repertoire.

Genes of the major histocompatibility complex (MHC) offer several assets that make them unique candidates for studies of adaptation in natural populations (Potts & Wakeland, 1990; Hedrick, 1994). The primary role of the MHC is to recognize foreign proteins, present them to specialist immune cells and initiate an immune response (Klein & Figueroa, 1986). The MHC gene family includes highly polymorphic genes encoding a set of transmembrane glycoproteins that are critical to the generation of immune responses (Kennedy et al., 2002). In general, foreign proteins enter cells either by infection or by phagocytosis into antigenpresenting cells such as macrophages. These foreign proteins are broken down into small peptides and loaded onto specific MHC molecules. The MHC molecule comprises an immunoglobulin stalk, which anchors the molecule to the cell surface, and a basket receptor called antigen-recognizing sites (ARS) located in peptide binding region (PBR). A subset of these protein/MHC complexes are then transported to the cell surface and presented for interrogation by the circulating T-cell population. A complex cascade of immune responses is triggered when the T cell binds to the presented peptide. Two major groups of MHC genes can be distinguished. MHC class I genes play an essential role in the immune defense against intracellular pathogens by binding peptides mainly derived from viral proteins and cancer-infected cells. They are expressed on the surface of all nucleated somatic cells. In contrast, MHC class II genes are predominantly involved in monitoring the extracellular environment by presenting peptides mainly derived from parasites (e.g. bacteria, nematodes, cestodes) to the T-cells. They are primarily expressed on antigen-presenting cells of the immune system, such as B cells and macrophages. Although ARS do show a degree of specificity, a single MHC molecule can bind multiple peptides that have common amino acids at particular anchor positions (Altuvia & Margalit, 2004). Genes within the MHC involved in antigen presentation constitute the most polymorphic loci known in vertebrates (Hedrick, 1994). The polymorphism of the MHC-molecules is associated with the diversity of the T-lymphocyte receptors that in turn determine the disease and parasite resistance of an organism and thus may affect the long-term survival rate of populations (Hedrick et al., 1999; Paterson et al., 1998). The ARS show high levels of polymorphism not only in the number of alleles but also in the sequence variation among alleles (Hughes & Yeager, 1998). The general view is that balancing selection is the determinant role in shaping patterns of nucleotide diversity in MHC genes (Bernatchez & Landry, 2003; Hughes & Nei, 1989).

Natural killer (NK) cells constitute an important branch of host innate immunity against tumors and viral infections. The activation of NK cells is controlled by the balance of activating and inhibitory receptors. Inhibitory NK receptors, such as members of Killer Immunoglobulin-like Receptors (KIR) and lectin-like CD94/NKG2 receptors, contain signaling ITIM motifs and recognize major histocompatibility-complex (MHC) antigens. Unlike MHC-recognizing T-cell antigen receptors, the inhibitory NK receptors are germline encoded and have fewer numbers compared to their MHC counterparts. They recognize self MHC molecules and exhibit allotype specificity instead of allele restriction. To investigate the mechanism of these largely nonpolymorphic receptors to recognize polymorphic MHC ligands, yet still retain their pathogen sensitivity, we have determined the crystal structures of human CD94 1], KIR2DL2 and its complex with a class I MHC ligand, HLA-Cw3 (Fig. 27.1a) [2,3]. Our results show that the inhibitory KIR receptor binds to the polymorphic α-helices of MHC in a similar mode as TCR.Unlike TCR, however, KIR receptors recognize conserved, rather than polymorphic residues, on MHC and have only minimum contacts with the eighth residue of the MHC-bound peptide, thus alleviatingthe peptide-MHC restriction of TCR.

B-cell epitope prediction is a computational approach originally developed to support the design of peptide-based vaccines for inducing protective antibody-mediated immunity, as exemplified by neutralization of biological activity (e.g., pathogen infectivity). Said approach is benchmarked against experimentally obtained data on paratope-epitope binding; but such data are curated primarily on the basis of immune-complex structure, obscuring the role of antigen conformational disorder in the underlying immune recognition process. This work aimed to critically analyze the curation of epitope-paratope binding data that are relevant to B-cell epitope prediction for peptide-based vaccine design. Database records on neutralizing monoclonal antipeptide antibody immune-complex structure were retrieved from the Immune Epitope Database (IEDB) and analyzed in relation to other data from both IEDB and external sources including the Protein Data Bank (PDB) and published literature, with special attention to data on conformational disorder among paratope-bound and unbound peptidic antigens. Data analysis revealed key examples of antipeptide antibodies that recognize conformationally disordered B-cell epitopes and thereby neutralize the biological activity of cognate targets (e.g., proteins and pathogens), with inconsistency noted in the mapping of some epitopes due to reliance on immune-complex structural details, which vary even among experiments utilizing the same paratope-epitope combination (e.g., with the epitope forming part of a peptide or a protein). The results suggest an alternative approach to curating paratope-epitope binding data based on neutralization of biological activity by polyclonal antipeptide antibodies, with reference to immunogenic peptide sequences and their conformational disorder in unbound antigen structures.

Human cytomegalovirus (HCMV) unique short region 3 (US3) protein, a type I membrane protein, prevents maturation of class I major histocompatibility complex (MHC) molecules by retaining them in the endoplasmic reticulum (ER) and thus helps inhibit antigen presentation to cytotoxic T cells. US3 molecules bind to class I MHC molecules in a transient fashion but retain them very efficiently in the ER nonetheless. The US3 luminal domain is responsible for ER retention of US3 itself, while both the US3 luminal and transmembrane domains are necessary for retaining class I MHC in the ER. We have expressed the luminal domain of US3 molecule in Escherichia coli and analyzed its secondary structure by using nuclear magnetic resonance. We then predicted the US3 tertiary structure by modeling it based on the US2 structure. Unlike the luminal domain of US2, the US3 luminal domain does not obviously interact with class I MHC molecules. The luminal domain of US3 dynamically oligomerizes in vitro and full-length US3 molecules associate with each other in vivo. We present a model depicting how dynamic oligomerization of US3 may enhance its ability to retain class I molecules within the ER.