Preconfiguration of the antigen-binding site during affinity maturation of a broadly neutralizing influenza virus antibody

Abstract

Affinity maturation refines a naive B-cell response by selecting mutations in antibody variable domains that enhance antigen binding. We describe a B-cell lineage expressing broadly neutralizing influenza virus antibodies derived from a subject immunized with the 2007 trivalent vaccine. The lineage comprises three mature antibodies, the unmutated common ancestor, and a common intermediate. Their heavy-chain complementarity determining region inserts into the conserved receptor-binding pocket of influenza HA. We show by analysis of structures, binding kinetics and long time-scale molecular dynamics simulations that antibody evolution in this lineage has rigidified the initially flexible heavy-chain complementarity determining region by two nearly independent pathways and that this preconfiguration accounts for most of the affinity gain. The results advance our understanding of strategies for developing more broadly effective influenza vaccines.immunity | antigen recognition | X-ray crystallography E xposure to a novel antigen, whether by infection or vaccination, induces an initial naive B-cell response. Cells bearing B-cell receptors (BCRs) that bind the antigen in question, even with relatively low affinity, proliferate selectively. In the continued presence of antigen, additional proliferation, accompanied by somatic hypermutation of the rearranged Ig heavy-and light-chain genes, leads to selection of cells with BCRs (and secreted antibodies) that bind more tightly to antigen than their precursors-a process known as affinity maturation (1-3). Recent methodological advances make it possible to study the history of this process in a given subject by isolating a number of individual B cells at a suitable time point after vaccination or infection and cloning their recombined heavy-and light-chain variable regions (4-6). If a subset of the variable regions thus identified derives from the same progenitor, one can infer the clonal lineage that gave rise to the observed genes, including the unmutated common ancestor (UCA) and the other unobserved intermediates at the interior nodes of the clonal tree with tips that are the genes of the mature antibodies (Fig. 1A). Structural and biochemical changes that occur during affinity maturation can be analyzed, and the mechanism of affinity enhancement elucidated.The influenza B-cell clonal lineage shown in Fig. 1A derives from plasmablasts sorted from a sample taken from an adult subject 1 wk after administration of the 2007 trivalent inactivated influenza virus vaccine. It includes just three mature B-cell clones. We have shown that one member of this lineage (CH65) bears a heavy-chain complementary determining region 3 (CDR H3) loop that inserts into the HA receptor-binding pocket, mimics the influenza virus receptor sialic acid, and has unusual breadth of neutralizing capacity (31 of 36 H1 strains tested) (7). We have now extended the structural and functional analysis to the entire lineage. By determining the structure and binding properties of the UCA and intermediate 2...

Overlapping hotspots in CDRs are critical sites for V region diversification

“…A similar analysis of other antigen selected and unselected human V regions should reveal why different V regions often dominate the response to different antigens and why we need so many V regions. Our findings also suggest that part of the process of developing new vaccine strategies should include the identification of motifs in those V regions that are likely to be initially be targeted AID and to use that information in planning which antigens should be used in sequential vaccines (1,30,(72)(73)(74). Similar genomewide analysis could also increase our understanding of the how AID is targeted to V and SRs and to certain off-target sites and contributes to our understanding of the biochemical mechanisms of these processes.…”

A common solution to group 2 influenza virus neutralization

“…In addition, access to a conserved epitope using different modes of binding and varied angles of approach is an emerging theme for glycan-dependent antibody recognition of HIV-1 Env (33) and has defined a supersite of vulnerability on HIV-1. This concept is emulated here for influenza virus, where stem antibodies approach the epitope in different ways and use distinct interactions but have functionally similar modes of neutralization, as also observed for bnAbs to the HA receptor binding site (9)(10)(11)(13)(14)(15).…”

“…Identification of antigenic sites on HA indicates that influenza antibodies are primarily directed against the immunodominant HA head region (7), which mediates endosomal uptake of the virus into host cells by binding to sialic acid receptors (8). Because of high mutation rates in the HA head region and its tolerance for antigenic changes, antibodies that target the HA head are typically only effective against strains closely related to the strain(s) by which they were elicited, although several receptor binding site-targeting antibodies with greater breadth have been structurally characterized (9)(10)(11)(12)(13)(14)(15). In contrast, antibodies that bind to the membrane-proximal HA stem region tend to exhibit much broader neutralizing activity and can target strains within entire subtypes and groups (16)(17)(18)(19)(20)(21)(22)(23)(24)(25) as well as across influenza types (24).…”

Characterization of pathogenic human monoclonal autoantibodies against GM-CSF

“…If another unrelated antigen initiates the antibody response and somatic mutations change the antigen-binding site so it binds to an epitope on GM-CSF, it is unlikely that chance somatic mutations would change the antigen-binding sites to bind four nonoverlapping, conformational epitopes on GM-CSF. Two crystallographic studies of affinity maturation of human antibodies (29,30) establish that the increase in affinity is due to stabilization of the original antigen contacts and that the antigen-binding site of the original antibody has the same general conformation as the affinity-matured, high-affinity antibodies. If subsequent affinity maturation is driven by GM-CSF, the epitope on the mutated antibody should still overlap the original epitope on GM-CSF.…”