Most antibodies induced by HIV-1 are ineffective at preventing initiation or spread of infection because they are either nonneutralizing or narrowly isolate-specific. Rare, ''broadly neutralizing'' antibodies have been detected that recognize relatively conserved regions on the envelope glycoprotein. Using stringently characterized, homogeneous preparations of trimeric HIV-1 envelope protein in relevant conformations, we have analyzed the molecular mechanism of neutralization by two of these antibodies, 2F5 and 4E10. We find that their epitopes, in the membrane-proximal segment of the envelope protein ectodomain, are exposed only on a form designed to mimic an intermediate state during viral entry. These results help explain the rarity of 2F5-and 4E10-like antibody responses and suggest a strategy for eliciting them. envelope glycoprotein ͉ membrane fusion H IV-1 infection generally induces a strong antibody response to the envelope glycoprotein [trimeric (gp160) 3 , cleaved to (gp120/gp41) 3 ], the sole antigen on the virion surface. Most induced antibodies are ineffective in preventing infection, however, because they are either nonneutralizing or narrowly isolate-specific, and the virus replicates so rapidly that ongoing selection of neutralization resistant mutants allows viral evolution to ''keep ahead'' of highaffinity antibody production (1). Moreover, much of the antibody response may be to rearranged or dissociated forms of gp120 and gp41, on which the dominant epitopes may be either in hypervariable loops or in positions occluded on virion-borne envelope trimer. Rare, ''broadly neutralizing'' antibodies have been detected that recognize one of three relatively conserved regions on the envelope protein: the CD4-binding site (mAb b12) (2); carbohydrates on the outer gp120 surface (mAb 2G12) (3); and a segment of the gp41 ectodomain adjacent to the viral membrane (mAbs 2F5 and 4E10) (4, 5), often called the ''membrane-proximal external region'' (MPER). We seek to understand the molecular mechanisms of neutralization by these and other antibodies.Fusion of viral and target-cell membranes initiates HIV-1 infection. Conformational changes in gp120 that accompany its binding to receptor (CD4) and coreceptor (e.g., CCR5 or CXCR4) lead to dissociation of gp120 from gp41 and a cascade of refolding events in the latter (6). In the course of these rearrangements, the N-terminal fusion peptide of gp41 translocates and inserts into the target-cell membrane. A proposed extended conformation of the gp41 ectodomain, with its fusion peptide thus inserted and the transmembrane anchor still in the viral membrane, has been called the ''prehairpin intermediate'' (7). It is the target of various fusion inhibitors, including T-20/enfuvirtide, the first approved fusioninhibiting antiviral drug (8), and the characteristics of the intermediate have been deduced from the properties of these inhibitors or mimicries by short gp41 fragments (9). Subsequent rearrangements from the intermediate to the postfusion state of gp41 involve ...

HIV-1 envelope spike (Env) is a type I membrane protein that mediates viral entry. We use NMR to determine an atomic structure of the transmembrane (TM) domain of HIV-1 Env reconstituted in bicelles that mimic a lipid bilayer. The TM forms a well-ordered trimer that protects a conserved membrane-embedded arginine. An N-terminal coiled-coil and a C-terminal hydrophilic core stabilize the trimer. Individual mutations of conserved residues did not disrupt the TM trimer and minimally affected membrane fusion and infectivity. Major changes in the hydrophilic core, however, altered the antibody sensitivity of Env. These results show how a TM domain anchors, stabilizes and modulates a viral envelope spike and suggest that its influence on Env conformation is an important consideration for HIV-1 immunogen design.

c Induction of broadly neutralizing antibodies (bNAbs) is an important goal for HIV-1 vaccine development. Two autoreactive bNAbs, 2F5 and 4E10, recognize a conserved region on the HIV-1 envelope glycoprotein gp41 adjacent to the viral membrane known as the membrane-proximal external region (MPER). They block viral infection by targeting a fusion-intermediate conformation of gp41, assisted by an additional interaction with the viral membrane. Another MPER-specific antibody, 10E8, has recently been reported to neutralize HIV-1 with potency and breadth much greater than those of 2F5 or 4E10, but it appeared not to bind phospholipids and might target the untriggered envelope spikes, raising the hope that the MPER could be harnessed for vaccine design without major immunological concerns. Here, we show by three independent approaches that 10E8 indeed binds lipid bilayers through two hydrophobic residues in its CDR H3 (third heavy-chain complementarity-determining region). Its weak affinity for membranes in general and preference for cholesterol-rich membranes may account for its great neutralization potency, as it is less likely than other MPER-specific antibodies to bind cellular membranes nonspecifically. 10E8 binds with high affinity to a construct mimicking the fusion intermediate of gp41 but fails to recognize the envelope trimers representing the untriggered conformation. Moreover, we can improve the potency of 4E10 without affecting its binding to gp41 by a modification of its lipid-interacting CDR H3. These results reveal a general mechanism of HIV-1 neutralization by MPER-specific antibodies that involves interactions with viral lipids.