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Molecular & Cellular Proteomics : MCP Mar 2022Major histocompatibility complex class II (MHC-II) antigen presentation underlies a wide range of immune responses in health and disease. However, how MHC-II antigen...
Major histocompatibility complex class II (MHC-II) antigen presentation underlies a wide range of immune responses in health and disease. However, how MHC-II antigen presentation is regulated by the peptide-loading catalyst HLA-DM (DM), its associated modulator, HLA-DO (DO), is incompletely understood. This is due largely to technical limitations: model antigen-presenting cell (APC) systems that express these MHC-II peptidome regulators at physiologically variable levels have not been described. Likewise, computational prediction tools that account for DO and DM activities are not presently available. To address these gaps, we created a panel of single MHC-II allele, HLA-DR4-expressing APC lines that cover a wide range of DO:DM ratio states. Using a combined immunopeptidomic and proteomic discovery strategy, we measured the effects DO:DM ratios have on peptide presentation by surveying over 10,000 unique DR4-presented peptides. The resulting data provide insight into peptide characteristics that influence their presentation with increasing DO:DM ratios. These include DM sensitivity, peptide abundance, binding affinity and motif, peptide length, and choice of binding register along the source protein. These findings have implications for designing improved HLA-II prediction algorithms and research strategies for dissecting the variety of functions that different APCs serve in the body.
Topics: Antigen Presentation; Antigen-Presenting Cells; Cell Line; HLA-D Antigens; HLA-DR Antigens; Histocompatibility Antigens Class II; Humans; Peptides; Proteomics
PubMed: 35085787
DOI: 10.1016/j.mcpro.2022.100204 -
Blood Nov 2020In this issue of , Margraf et al selectively delete integrin linked kinase (ILK) in myeloid cells of mice to show that this integrin-binding protein suppresses...
In this issue of , Margraf et al selectively delete integrin linked kinase (ILK) in myeloid cells of mice to show that this integrin-binding protein suppresses chemokine-induced neutrophil extravasation and ischemia-induced reperfusion injury.
Topics: CD18 Antigens; Chemokines; Humans; Inflammation; Lymphocyte Function-Associated Antigen-1; Neutrophils; Protein Serine-Threonine Kinases
PubMed: 33152089
DOI: 10.1182/blood.2020008316 -
PLoS Pathogens Oct 2023Neutralizing antibodies (NAbs) to multiple epitopes on the HIV-1-envelope glycoprotein (Env) have been isolated from infected persons. The potency of NAbs is measured...
Neutralizing antibodies (NAbs) to multiple epitopes on the HIV-1-envelope glycoprotein (Env) have been isolated from infected persons. The potency of NAbs is measured more often than the size of the persistent fraction of infectivity at maximum neutralization, which may also influence preventive efficacy of active or passive immunization and the therapeutic outcome of the latter. Many NAbs neutralize HIV-1 CZA97.012, a clone of a Clade-C isolate, to ~100%. But here NAb PGT151, directed to a fusion-peptide epitope, left a persistent fraction of 15%. NAb PGT145, ligating the Env-trimer apex, left no detectable persistent fraction. The divergence in persistent fractions was further analyzed by depletion of pseudoviral populations of the most PGT151- and PGT145-reactive virions. Thereby, neutralization by the non-depleting NAb increased, whereas neutralization by the depleting NAb decreased. Furthermore, depletion by PGT151 increased sensitivity to autologous neutralization by sera from rabbits immunized with soluble native-like CZA97.012 trimer: substantial persistent fractions were reduced. NAbs in these sera target epitopes comprising residue D411 at the V4-β19 transition in a defect of the glycan shield on CZA97.012 Env. NAb binding to affinity-fractionated soluble native-like CZA97.012 trimer differed commensurately with neutralization in analyses by ELISA and surface plasmon resonance. Glycan differences between PGT151- and PGT145-purified trimer fractions were then demonstrated by mass spectrometry, providing one explanation for the differential antigenicity. These differences were interpreted in relation to a new structure at 3.4-Å resolution of the soluble CZA97.012 trimer determined by cryo-electron microscopy. The trimer adopted a closed conformation, refuting apex opening as the cause of reduced PGT145 binding to the PGT151-purified form. The evidence suggests that differences in binding and neutralization after trimer purification or pseudovirus depletion with PGT145 or PGT151 are caused by variation in glycosylation, and that some glycan variants affect antigenicity through direct effects on antibody contacts, whereas others act allosterically.
Topics: Animals; Rabbits; HIV Antibodies; HIV-1; Cryoelectron Microscopy; Antibodies, Neutralizing; Epitopes; Antigens, Viral; Polysaccharides; env Gene Products, Human Immunodeficiency Virus; HIV Infections
PubMed: 37903160
DOI: 10.1371/journal.ppat.1011601 -
Chembiochem : a European Journal of... Sep 2022Antibodies recognize their cognate antigens with high affinity and specificity, but the prediction of binding sites on the antigen (epitope) corresponding to a specific...
Antibodies recognize their cognate antigens with high affinity and specificity, but the prediction of binding sites on the antigen (epitope) corresponding to a specific antibody remains a challenging problem. To address this problem, we developed AbAdapt, a pipeline that integrates antibody and antigen structural modeling with rigid docking in order to derive antibody-antigen specific features for epitope prediction. In this study, we systematically assessed the impact of integrating the state-of-the-art protein modeling method AlphaFold with the AbAdapt pipeline. By incorporating more accurate antibody models, we observed improvement in docking, paratope prediction, and prediction of antibody-specific epitopes. We further applied AbAdapt-AF in an anti-receptor binding domain (RBD) antibody complex benchmark and found AbAdapt-AF outperformed three alternative docking methods. Also, AbAdapt-AF demonstrated higher epitope prediction accuracy than other tested epitope prediction tools in the anti-RBD antibody complex benchmark. We anticipate that AbAdapt-AF will facilitate prediction of antigen-antibody interactions in a wide range of applications.
Topics: Antibodies; Antibody Specificity; Antigens; Binding Sites, Antibody; Epitopes
PubMed: 35893479
DOI: 10.1002/cbic.202200303 -
Journal of Molecular Graphics &... Jan 2023Specific antibodies can bind to protein antigens with high affinity and specificity, and this property makes them one of the best protein-based therapeutics. Accurate...
Specific antibodies can bind to protein antigens with high affinity and specificity, and this property makes them one of the best protein-based therapeutics. Accurate prediction of antibody‒protein antigen binding affinity is crucial for designing effective antibodies. The current predictive methods for protein‒protein binding affinity usually fail to predict the binding affinity of an antibody‒protein antigen complex with a comparable level of accuracy. Here, new models specific for antibody‒antigen binding affinity prediction are developed according to the different types of interface and surface areas present in antibody‒antigen complex. The contacts-based descriptors are also employed to construct or train different models specific for antibody‒protein antigen binding affinity prediction. The results of this study show that (i) the area-based descriptors are slightly better than the contacts-based descriptors in terms of the predictive power; (ii) the new models specific for antibody‒protein antigen binding affinity prediction are superior to the previously-used general models for predicting the protein‒protein binding affinities; (iii) the performances of the best area-based and contacts-based models developed in this work are better than the performances of a recently-developed graph-based model (i.e., CSM-AB) specific for antibody‒protein antigen binding affinity prediction. The new models developed in this work would not only help understand the mechanisms underlying antibody‒protein antigen interactions, but would also be of some applicable utility in the design and virtual screening of antibody-based therapeutics.
Topics: Proteins; Protein Binding; Antigen-Antibody Complex; Machine Learning; Antigens
PubMed: 36356467
DOI: 10.1016/j.jmgm.2022.108364 -
FEBS Letters Oct 2022Mtb8.4, a secretory T-cell antigen of Mycobacterium tuberculosis, is important for providing an antigen-specific immune response. In this study, we showed Mtb8.4 to have...
Mtb8.4, a secretory T-cell antigen of Mycobacterium tuberculosis, is important for providing an antigen-specific immune response. In this study, we showed Mtb8.4 to have both heme-binding and fibril-forming properties, using experimental and in silico methods. High absorbance at 410 nm and interaction with hemin-agarose demonstrated its heme-binding nature. Titration of Mtb8.4 with heme resulted in 1 : 1 stoichiometry. The heme-binding pocket in Mtb8.4 was identified by molecular modeling, and binding residues were predicted using molecular docking. The molecular dynamics simulations of apo- and heme-bound Mtb8.4 confirmed that the heme group forms a stable complex. Transmission electron microscopy analyses and dye-binding assays showed that Mtb8.4 forms fibers. Computational studies predicted that the C-terminal sequence ( AAQYIGLVESV ) is important for forming fibers. In silico analyses further anticipated the probable epitope ( AMAAQLQAV ) of Mtb8.4. The fiber-forming properties of Mtb8.4 could be advantageous from a vaccine perspective for aggregate/fibril-based vaccine delivery or it might influence the epitope presentation of Mtb8.4.
Topics: Mycobacterium tuberculosis; Antigens, Bacterial; Molecular Weight; Molecular Docking Simulation; Heme; T-Lymphocytes; Epitopes
PubMed: 35795993
DOI: 10.1002/1873-3468.14446 -
FEBS Letters Aug 2020Factor H binding protein (fHbp) is a key virulence factor of Neisseria meningitidis and a main component of the two licensed vaccines against serogroup B meningococcus... (Review)
Review
Factor H binding protein (fHbp) is a key virulence factor of Neisseria meningitidis and a main component of the two licensed vaccines against serogroup B meningococcus (Bexsero and Trumenba). fHbp is a surface-exposed lipoprotein that enables the bacterium to survive in human blood by binding the human complement regulator factor H (fH). When used as vaccine, the protein induces antibodies with potent bactericidal activity. While the fHbp gene is present in the majority of N. meningitidis serogroup B isolates, the expression level varies up to 15 times between different strains and more than 700 different sequence variants have been described. Antigenically, the protein has been divided into three variants or two subfamilies. The 3D structure of fHbp alone, in combination with fH or in complex with bactericidal antibodies, has been key to understanding the molecular details of the protein. In this article, we will review the biochemical and immunological properties of fHbp, and its key role in meningococcal pathogenesis, complement regulation, and immune evasion.
Topics: Animals; Antigens, Bacterial; Bacterial Proteins; Complement Factor H; Gene Expression Regulation, Bacterial; Humans; Immune Evasion; Meningococcal Infections; Meningococcal Vaccines; Neisseria meningitidis, Serogroup B; Protein Domains
PubMed: 32298465
DOI: 10.1002/1873-3468.13793 -
Genome Medicine Aug 2019Neoantigens are newly formed peptides created from somatic mutations that are capable of inducing tumor-specific T cell recognition. Recently, researchers and clinicians... (Review)
Review
Neoantigens are newly formed peptides created from somatic mutations that are capable of inducing tumor-specific T cell recognition. Recently, researchers and clinicians have leveraged next generation sequencing technologies to identify neoantigens and to create personalized immunotherapies for cancer treatment. To create a personalized cancer vaccine, neoantigens must be computationally predicted from matched tumor-normal sequencing data, and then ranked according to their predicted capability in stimulating a T cell response. This candidate neoantigen prediction process involves multiple steps, including somatic mutation identification, HLA typing, peptide processing, and peptide-MHC binding prediction. The general workflow has been utilized for many preclinical and clinical trials, but there is no current consensus approach and few established best practices. In this article, we review recent discoveries, summarize the available computational tools, and provide analysis considerations for each step, including neoantigen prediction, prioritization, delivery, and validation methods. In addition to reviewing the current state of neoantigen analysis, we provide practical guidance, specific recommendations, and extensive discussion of critical concepts and points of confusion in the practice of neoantigen characterization for clinical use. Finally, we outline necessary areas of development, including the need to improve HLA class II typing accuracy, to expand software support for diverse neoantigen sources, and to incorporate clinical response data to improve neoantigen prediction algorithms. The ultimate goal of neoantigen characterization workflows is to create personalized vaccines that improve patient outcomes in diverse cancer types.
Topics: Antigen Presentation; Antigens, Neoplasm; Cancer Vaccines; Computational Biology; Gene Expression; HLA Antigens; Histocompatibility Testing; Humans; Mutation; Neoplasms; Peptides; Protein Binding; Receptors, Antigen, T-Cell; T-Lymphocytes; Workflow
PubMed: 31462330
DOI: 10.1186/s13073-019-0666-2 -
Immunology and Cell Biology Feb 2022MHC-related protein 1 (MR1) presents microbial riboflavin metabolites to mucosal-associated invariant T (MAIT) cells for surveillance of microbial presence. MAIT cells...
MHC-related protein 1 (MR1) presents microbial riboflavin metabolites to mucosal-associated invariant T (MAIT) cells for surveillance of microbial presence. MAIT cells express a semi-invariant T-cell receptor (TCR), which recognizes MR1-antigen complexes in a pattern-recognition-like manner. Recently, diverse populations of MR1-restricted T cells have been described that exhibit broad recognition of tumor cells and appear to recognize MR1 in association with tumor-derived self-antigens, though the identity of these antigens remains unclear. Here, we have used TCR gene transfer and engineered MR1-expressing antigen-presenting cells to probe the MR1 restriction and antigen reactivity of a range of MR1-restricted TCRs, including model tumor-reactive TCRs. We confirm MR1 reactivity by these TCRs, show differential dependence on lysine at position 43 of MR1 (K43) and demonstrate competitive inhibition by the MR1 ligand 6-formylpterin. TCR-expressing reporter lines, however, failed to recapitulate the robust tumor specificity previously reported, suggesting an importance of accessory molecules for MR1-dependent tumor reactivity. Finally, MR1-mutant cell lines showed that distinct residues on the α1/α2 helices were required for TCR binding by different MR1-restricted T cells and suggested central but distinct docking modes by the broad family of MR1-restricted αβ TCRs. Collectively, these data are consistent with recognition of distinct antigens by diverse MR1-restricted T cells.
Topics: Histocompatibility Antigens Class I; Minor Histocompatibility Antigens; Mucosal-Associated Invariant T Cells; Receptors, Antigen, T-Cell; Receptors, Antigen, T-Cell, alpha-beta
PubMed: 34940995
DOI: 10.1111/imcb.12519 -
Biochimica Et Biophysica Acta.... Nov 2021Protective antigen channel is the central component of the deadly anthrax exotoxin responsible for binding and delivery of the toxin's enzymatic lethal and edema factor... (Review)
Review
Protective antigen channel is the central component of the deadly anthrax exotoxin responsible for binding and delivery of the toxin's enzymatic lethal and edema factor components into the cytosol. The channel, which is more than three times longer than the lipid bilayer membrane thickness and has a 6-Å limiting diameter, is believed to provide a sophisticated unfoldase and translocase machinery for the foreign protein transport into the host cell cytosol. The tripartite toxin can be reengineered, one component at a time or collectively, to adapt it for the targeted cancer therapeutic treatments. In this review, we focus on the biophysical studies of the protective antigen channel-forming activity, small ion transport properties, enzymatic factor translocation, and blockage comparing it with the related clostridial binary toxin channels. We address issues linked to the anthrax toxin channel structural dynamics and lipid dependence, which are yet to become generally recognized as parts of the toxin translocation machinery.
Topics: Antigens, Bacterial; Bacterial Toxins; Hydrogen-Ion Concentration; Lipid Bilayers; Protein Conformation; Protein Transport
PubMed: 34332985
DOI: 10.1016/j.bbamem.2021.183715