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HIV-1 entry into cells is mediated by the envelope glycoprotein receptor-binding (gp120) and membrane fusion-promoting (gp41) subunits. The gp41 heptad repeat 1 (HR1) domain is the molecular target of the fusion-inhibitor drug enfuvirtide (T20). The HR1 sequence is highly conserved and therefore considered an attractive target for vaccine development, but it is unknown whether antibodies can access HR1. Herein, we use gp41-based peptides to select a human antibody, 5H/I1-BMV-D5 (D5), that binds to HR1 and inhibits the assembly of fusion intermediates in vitro . D5 inhibits the replication of diverse HIV-1 clinical isolates and therefore represents a previously unknown example of a crossneutralizing IgG selected by binding to designed antigens. NMR studies and functional analyses map the D5-binding site to a previously identified hydrophobic pocket situated in the HR1 groove. This hydrophobic pocket was proposed as a drug target and subsequently identified as a common binding site for peptide and peptidomimetic fusion inhibitors. The finding that the D5 fusion-inhibitory antibody shares the same binding site suggests that the hydrophobic pocket is a “hot spot” for fusion inhibition and an ideal target on which to focus a vaccine-elicited antibody response. Our data provide a structural framework for the design of new immunogens and therapeutic antibodies with crossneutralizing potential.

The causative agent of a recent outbreak of an atypical pneumonia, known as severe acute respiratory syndrome (SARS), has been identified as a coronavirus (CoV) not belonging to any of the previously identified groups. Fusion of coronaviruses with the host cell is mediated by the envelope spike protein. Two regions within the spike protein of SARS-CoV have been identified, showing a high degree of sequence conservation with the other CoV, which are characterized by the presence of heptad repeats (HR1 and HR2). By using synthetic and recombinant peptides corresponding to the HR1 and HR2 regions, we were able to characterize the fusion-active complex formed by this novel CoV by CD, native PAGE, proteolysis protection analysis, and size-exclusion chromatography. HR1 and HR2 of SARS-CoV associate into an antiparallel six-helix bundle, with structural features typical of the other known class I fusion proteins. We have also mapped the specific boundaries of the region, within the longer HR1 domain, making contact with the shorter HR2 domain. Notably, the inner HR1 coiled coil is a stable α-helical domain even in the absence of interaction with the HR2 region. Inhibitors binding to HR regions of fusion proteins have been shown to be efficacious against many viruses, notably HIV. Our results may help in the design of anti-SARS therapeutics.

The HIV-1 gp41 envelope glycoprotein promotes fusion of the virus and cell membranes through the formation of a trimer-of-hairpins structure, in which the amino- and carboxyl-terminal regions of the gp41 ectodomain are brought together. Synthetic peptides derived from these two regions (called N and C peptides, respectively) inhibit HIV-1 entry. In contrast to C peptides, which inhibit in the nanomolar range, N peptides are weak inhibitors with IC 50 values in the micromolar range. To test the hypothesis that the weak inhibition of N peptides results from their tendency to aggregate, we have constructed chimeric variants of the N-peptide region of gp41 in which soluble trimeric coiled coils are fused to portions of the gp41 N peptide. These molecules, which present the N peptide in a trimeric coiled-coil conformation, are remarkably more potent inhibitors than the N peptides themselves and likely target the carboxyl-terminal region of the gp41 ectodomain. The best inhibitors described here inhibit HIV-1 entry at nanomolar concentrations.

Mitochondrial fission is mediated by the dynamin-related GTPases Dnm1/Drp1 (yeast/mammals), which form spirals around constricted sites on mitochondria. Additional membrane-associated adaptor proteins (Fis1, Mdv1, Mff, and MiDs) are required to recruit these GTPases from the cytoplasm to the mitochondrial surface. Whether these adaptors participate in both GTPase recruitment and membrane scission is not known. Here we use a yeast strain lacking all fission proteins to identify the minimal combinations of GTPases and adaptors sufficient for mitochondrial fission. Although Fis1 is dispensable for fission, membrane-anchored Mdv1, Mff, or MiDs paired individually with their respective GTPases are sufficient to divide mitochondria. In addition to their role in Drp1 membrane recruitment, MiDs coassemble with Drp1 in vitro. The resulting heteropolymer adopts a dramatically different structure with a narrower diameter than Drp1 homopolymers assembled in isolation. This result demonstrates that an adaptor protein alters the architecture of a mitochondrial dynamin GTPase polymer in a manner that could facilitate membrane constriction and severing activity.

Influenza A virus causes seasonal flu epidemics and periodic worldwide pandemics (e.g., the 1918 Spanish flu, which caused ≈50 million deaths). The viral surface protein HA is the primary target of neutralizing Abs in natural infections (1). At any given time there are a limited number of viral strains circulating in humans, restricting widespread immunity to a small subset of potential viruses. Yearly seasonal epidemics arise from antigenic drift in the sequence of HA of currently circulating viruses, whereas pandemics are caused by the emergence of new, antigenically divergent viruses to which there is little to no immunity in the population (i.e., antigenic shift). The worldwide spread of a new H1N1 virus in 2009 caused the first recorded pandemic in more than 40 years. Current influenza vaccines are primarily produced from killed virus and mimic natural infection, inducing strain-specific, mainly HA-based, neutralization. Vaccine is produced from representative circulating strains grown in chicken eggs in a months-long process. Recent efforts aim to produce a broader influenza vaccine that focuses on common neutralization epitopes shared by multiple influenza strains. Such a vaccine should target a variety of influenza strains and better combat pandemics. A study published in PNAS describes an exciting strategy toward developing a broader influenza vaccine (2).