Identification of autoantigens recognized by the 2F5 and 4E10 broadly neutralizing HIV-1 antibodies.
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2013-02-11
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Abstract
Many human monoclonal antibodies that neutralize multiple clades of HIV-1 are polyreactive and bind avidly to mammalian autoantigens. Indeed, the generation of neutralizing antibodies to the 2F5 and 4E10 epitopes of HIV-1 gp41 in man may be proscribed by immune tolerance because mice expressing the V(H) and V(L) regions of 2F5 have a block in B cell development that is characteristic of central tolerance. This developmental blockade implies the presence of tolerizing autoantigens that are mimicked by the membrane-proximal external region of HIV-1 gp41. We identify human kynureninase (KYNU) and splicing factor 3b subunit 3 (SF3B3) as the primary conserved, vertebrate self-antigens recognized by the 2F5 and 4E10 antibodies, respectively. 2F5 binds the H4 domain of KYNU which contains the complete 2F5 linear epitope (ELDKWA). 4E10 recognizes an epitope of SF3B3 that is strongly dependent on hydrophobic interactions. Opossums carry a rare KYNU H4 domain that abolishes 2F5 binding, but they retain the SF3B3 4E10 epitope. Immunization of opossums with HIV-1 gp140 induced extraordinary titers of serum antibody to the 2F5 ELDKWA epitope but little or nothing to the 4E10 determinant. Identification of structural motifs shared by vertebrates and HIV-1 provides direct evidence that immunological tolerance can impair humoral responses to HIV-1.
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Yang, Guang, T Matt Holl, Yang Liu, Yi Li, Xiaozhi Lu, Nathan I Nicely, Thomas B Kepler, S Munir Alam, et al. (2013). Identification of autoantigens recognized by the 2F5 and 4E10 broadly neutralizing HIV-1 antibodies. J Exp Med, 210(2). pp. 241–256. 10.1084/jem.20121977 Retrieved from https://hdl.handle.net/10161/10900.
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Scholars@Duke
S. Munir Alam
Research Interests.
The Alam laboratory’s primary research is focused on understanding the biophysical properties of antigen-antibody binding and the molecular events of early B cell activation using the HIV-1 broadly neutralizing antibody (bnAb) lineage models. We are studying how HIV-1 Envelope proteins of varying affinities are sensed by B cells expressing HIV-1 bnAbs or their germline antigen receptors and initiate early signaling events for their activation. In the long-term these studies will facilitate design and pre-selection of immunogens for testing in animal models and accelerate HIV-1 vaccine development.
Current research include the following NIAID-funded projects
Antigen recognition and activation of B cell antigen receptors with the specificity of HIV-1 broadly neutralizing antibodies. This project involves elucidating the early events on the B cell surface following antigen (Ag) engagement of the B cell antigen receptor (BCR) and to provide an assessment of the in vivo potential of an Ag to drive B cell activation. We are performing biophysical interactions analyses and using high-resolution microscopy to define the physico-chemical properties of BCR-Ag interactions that govern signaling and activation thresholds for BCR triggering and the BCR endocytic function in antigen internalization. The overall objective of these studies is to bridge the quantitative biophysical and membrane dynamics measurements of Ag-BCR interactions to ex-vivo and in-vivo B cell activation. This NIAID-funded research is a collaboration with co-investigators Professor Michael Reth (University of Freiburg, Germany) and Dr. Laurent Verkoczy (San Diego Biomedical Research Institute, CA).
Immunogen Design for Induction of HIV gp41 Broadly Neutralizing Antibodies. This research project addresses the critical problem of vaccine induction of disfavored HIV-1 antibody lineages, like those that target the membrane proximal external region (MPER) of HIV Env gp41. This program combines structure and lineage-based vaccine development strategies to design immunogens that will induce bnAb lineages that are not polyreactive and therefore easier to induce. The overall objective of this program grant is to develop and test sequential immunogens that will initiate and induce HIV-1 bnAb lineages like the potent MPER bnAb DH511. Using a germline-targeting (GT) epitope scaffold design and a prime/boost strategy, we are testing induction of DH511-like bnAbs in knock-in (KI) mice models expressing the DH511 germline receptors. This P01 research program is in collaboration with Dr. William Schief (The Scripps Research Institute, CA), who leads the team that are designing germline targeting (GT)-scaffold prime and boost immunogens and Dr. Ming Tian at Harvard University who developed relevant knock-mice models for the study.
Derek Wilson Cain
My research focuses on the interactions of T cells and B cells during infection or following vaccination. I am particularly interested in the inter- and intracellular events that take place within germinal centers, the anatomic site of antibody evolution during an immune response.
Leonard D. Spicer
The focus of this laboratory is the study of structure/function relationships in biological macromolecules and their binding interactions. The principal method we use for system characterization is magnetic resonance spectroscopy. One specific area of interest is the structural characterization of functional domains in proteins which regulate the transcription of DNA coding for biosynthetic enzymes. The system under current investigation is the methionine repressor protein metJ, its corepressor S-adenosylmethionine, and the cognate sequence DNA. This protein, which functions as a dimer, exhibits a recently described DNA binding motif involving insertion of two beta strands into the major groove with additional stabilization of the complex arising from helix contacts at the dimer-dimer interface. We are using a full complement of heteronuclear 3D and 4D NMR methods to aid in the assignment of the main chain of the metJ repressor. We have recently reported a thermodynamic analysis of the binding interactions of metJ with its cognate DNA and corepressor SAM. We are now developing methods to measure fast proton exchange rates to complement our planned solution structural characterization. We have just initiated another project in collaboration with scientists at the Pacific Northwest National Laboratory to study macromelecular structures of DNA repair proteins in the nucleotide excision repair pathway. The first components of this critical supramacromolecular assembly we are investigating involve the DNA binding domain of the XPA protein for which we are determining the global fold in solution by NMR. Our program also includes a systematic approach to characterizing the conformational preferences of a number of sequentially related peptides developed by Dr. Barton Haynes' laboratory as candidate vaccines for HIV. The peptides consist of a fusion of two noncontiguous segments of the HIV protein gp120. Our goal is to establish whether structural conformers in solution contribute to peptide immunogenicity. We have finished a careful conformational analysis of the initial four peptides and are now correlating the conformer similarities and differences with immunogenic properties. We have also rationally designed several new peptides based on structural criteria and corresponding structural homology to the heavy fragment of IgA proteins. Initial NMR analysis and immunogenic response to three of the designed mutants indicate the rational design of preferred conformers was successful, but raised some novel questions regarding function of immunogenic peptides. We have also just begun a study of solution conformations of the hypoglycosylated tumor specific epitope repeat unit of human mucin and a promising mutant identified by Dombrowski and Wright. This epitope is common to breast and other adenocarcinomas and regulation of tumor specific lymphoid cells responding to this immunogen may be an important step in tumor control. Another protein under investigation is a functional core packing mutant of thioredoxin. We have fully characterized backbone chain dynamics to assess the impact of this mutation on molecular motions and are currently determining its high resolution tertiary structure. Currently, we are also using this mutant to demonstrate a new approach to global fold determination using a minimum set of long range NMR constraints. Finally, as an essential part of these studies, we are developing and have reported new 3- and 4-dimensional NMR experiments and heteronuclear filters for application to large protein systems and binding complexes.
Finally, the core activities of the NMR Center staff have continued to progress rapidly and enhancements to the state-of-the-art instrumentation have again been incorporated. A new deuteration strategy for assignment and study of large proteins by NMR has been developed and used to characterize one of the largest protein monomer reported to date, human carbonic anhydrase. We have also shown that we can observe the longest range distance constraints to date from NOESY correlations which are important in determining tertiary structure of proteins and we are examining the efficacy of structure determinations based on using these critical but limited constraints.
Garnett H. Kelsoe
- Lymphocyte development and antigen-driven diversification of immunoglobulin and T cell antigen receptor genes.
2. The germinal center reaction and mechanisms for clonal selection and self - tolerance. The origins of autoimmunity.
3. Interaction of innate- and adaptive immunity and the role of inflammation in lymphoid organogenesis.
4. The role of secondary V(D)J gene rearrangment in lymphocyte development and malignancies.
5. Mathematical modeling of immune responses, DNA motifs, collaborations in bioinformatics.
6. Humoral immunity to influenza and HIV-1.
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