172 research outputs found

    Ag binding sites coverage by consensus and method-specific residues.

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    <p>For each set, we recorded the total number of residues, the number of Ag contacting residues and the percentage of Ag binding sites coverage. In all of the comparisons, Paratome-specific residues covered a significantly larger portion of the Ag binding sites.</p

    Structural Consensus among Antibodies Defines the Antigen Binding Site

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    <div><p>The Complementarity Determining Regions (CDRs) of antibodies are assumed to account for the antigen recognition and binding and thus to contain also the antigen binding site. CDRs are typically discerned by searching for regions that are most different, in sequence or in structure, between different antibodies. Here, we show that ∼20% of the antibody residues that actually bind the antigen fall outside the CDRs. However, virtually all antigen binding residues lie in regions of structural consensus across antibodies. Furthermore, we show that these regions of structural consensus which cover the antigen binding site are identifiable from the sequence of the antibody. Analyzing the predicted contribution of antigen binding residues to the stability of the antibody-antigen complex, we show that residues that fall outside of the traditionally defined CDRs are at least as important to antigen binding as residues within the CDRs, and in some cases, they are even more important energetically. Furthermore, antigen binding residues that fall outside of the structural consensus regions but within traditionally defined CDRs show a marginal energetic contribution to antigen binding. These findings allow for systematic and comprehensive identification of antigen binding sites, which can improve the understanding of antigenic interactions and may be useful in antibody engineering and B-cell epitope identification.</p> </div

    Automated ABRs Identification

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    <p>(<b>A</b>) <b>Sequence based ABRs identification.</b> A BLAST search is performed using the query Ab sequence versus the dataset of non-redundant PDB Abs. Using the best hit from the BLAST search, the query and annotated Abs FRs are aligned and hence the query sequence ABRs are inferred based on the location of the annotated sequence ABRs in the MSTA. (<b>B</b>) <b>Structure based ABRs identification.</b> A BLAST search is performed using the sequence of the query Ab versus our dataset of Abs. Using the best hit from the BLAST search, the query and annotated Abs are structurally aligned. The ABRs of the query Ab are inferred based on the location of the annotated Ab ABRs in the MSTA.</p

    Contribution of Paratome-unique and CDR-unique residues to the binding energy in Ab-Ag complexes.

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    <p>(A) The distributions of ΔΔG values of an in-silico alanine scan analysis of Paratome-unique, CDRs-unique and CDR Ag binding residues. ΔΔG values ranging between −0.25 and 0.25 were defined as neutral. ΔΔG values<−0.25 were defined as stabilizing. ΔΔG values >0.25 were defined as destabilizing. It is clear from the distributions that typically, a Paratome-unique residue is at least as energetically important as a residue in the CDRs, while a CDR unique residue is less energetically important relative to residues within the CDRs that are identified by Paratome. (B)+(C) A detailed analysis of anti-IL-15 Ab with human IL-15 (PDB 2xqb). (B) The surface of the Ag chain is rendered according to atom charge. Due to the hydrogen bond with antigenic GLU53, TYR49 is located in high proximity to the Ag. Ab LEU46 is located in proximity to antigenic LEU52. (C) Seven residues from L2 (green, solid ribbon) interact with the Ag (orange, solid ribbon). Two of them (LEU46 and TYR49) are not identified as part of the CDR by any other CDR identification method. These Paratome-unique residues and the antigenic residues they contact (LEU52 and GLU53) are depicted in sticks. LEU46 forms a hydrophobic interaction with LEU52. TYR49 forms a hydrogen bond with antigenic GLU53 as well as a cation- п interaction with ARG53 of the Ab. Both contribute substantially to Ag binding (see text).</p

    Recall and precision of Ag binding sites identification.

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    <p>Average precision and recall were calculated for the Abs in the test set for Paratome, Kabat, Chothia and IMGT methods. Error bars represent standard error of the mean.</p

    Total number of residues identified by each method for all Ab-Ag complexes in the test set.

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    <p>L1–L3 are ABR/CDR1-3 of the light chain. H1–H3 are ABR/CDR1-3 of the heavy chain. Total light and heavy are the sum of all identified residues in the light and heavy chains respectively.</p

    Structure-based identification of ABRs.

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    <p>(A) Using the non-redundant set of all Ab-Ag complexes in the PDB, (B) we created a multiple structure alignment of the Abs. Residues that are in contact with the Ag were identified by searching for structurally aligned positions that systematically create contacts with the Ag (black and grey solid circles) and disregarded positions that contact the Ag only sporadically (open shapes). (C) The contacting positions were mapped to the sequence representation of the multiple structure alignment (bold letters). The stretches of amino acids in which at least 10% of the Abs are in contact with the Ag were defined as ABRs (white rectangle).</p

    Average Ag binding sites recall and precision of light and heavy chains for all ABRs/CDRs.

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    <p>(A) Average Ag binding sites precision (B) Average Ag binding sites recall. Error bars represent standard error of the mean.</p

    ImmunomeBrowser analysis of B and T cell epitopes.

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    <p>R/T, number responded over number tested; RFscore, response frequency score; N-arm, N-terminal arm; NTD, N-terminal domain; CP, cytoplasmic, EC, extracellular; HBD, heparin binding domain; HRA/HRB, heptad repeats; DI, docking inhibition; Cys-noose, cysteine noose; bovine host*, natural infection in children/infants**, natural infection in adults***; bold indicates overlap.</p><p>ImmunomeBrowser analysis of B and T cell epitopes.</p

    Analysis of Human RSV Immunity at the Molecular Level: Learning from the Past and Present

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    <div><p>Human RSV is one of the most prevalent viral pathogens of early childhood for which no vaccine is available. Herein we provide an analysis of RSV epitope data to examine its application to vaccine design and development. Our objective was to provide an overview of antigenic coverage, identify critical antibody and T cell determinants, and then analyze the cumulative RSV epitope data from the standpoint of functional responses using a combinational approach to characterize antigenic structure and epitope location. A review of the cumulative data revealed, not surprisingly, that the vast majority of epitopes have been defined for the two major surface antigens, F and G. Antibody and T cell determinants have been reported from multiple hosts, including those from human subjects following natural infection, however human data represent a minority of the data. A structural analysis of the major surface antigen, F, showed that the majority of epitopes defined for functional antibodies (neutralizing and/or protective) were either shown to bind pre-F or to be accessible in both pre- and post-F forms. This finding may have has implications for on-going vaccine design and development. These interpretations are in agreement with previous work and can be applied in the larger context of functional epitopes on the F protein. It is our hope that this work will provide the basis for further RSV-specific epitope discovery and investigation into the nature of antigen conformation in immunogenicity.</p></div
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