Adjacent Trimers Research Articles

Distinct binding mode of BAFF antagonist antibodies belimumab and tabalumab, analyzed by computer simulation.

B cell-activating factor (BAFF) can bind with specific receptors to activate signalling pathways associated with the B cell activation. Belimumab and tabalumab are anti-BAFF (B cell depleting) monoclonal antibodies, with therapeutic efficacy demonstrated for the treatment of autoimmune disorders, while belimumab was approved by FDA in 2011 as a targeted therapy for systemic lupus erythematosus (SLE) and exhibited better clinical outcome than tabalumab. In this investigation, the combination modes of BAFF-belimumab and BAFF-tabalumab complexes were simulated in silico to better understand the reason for the comparative inhibitory difference between belimumab and tabalumab. The structures of belimumab and tabalumab were constructed through homology modelling. The combination mode of BAFF-belimumab complex was analyzed by molecular dynamics simulation, while that of BAFF-tabalumab complex was analyzed by protein-protein docking following the molecular dynamics simulation. Both belimumab and tabalumab were bound with BAFF at the same hydrophobic center to which the natural receptors of BAFF bind as well. Belimumab heavy chain components I51, F54, K58, D100, D101, L102, L103, and P105 and R27, Y30, K49, and S65 of belimumab light chain contribute to the BAFF-belimumab interaction mainly via hydrogen bonds, salt bridges, and hydrophobic interactions. More importantly, belimumab could bind to L83 of BAFF and produce steric hindrance with the adjacent BAFF trimers, while tabalumab could not. Therefore, our results indicated that belimumab has a better clinical outcome compared with tabalumab mainly because belimumab could bind to L83 of BAFF and interfere the formation of a BAFF 60-mer, besides mediating inhibition of the interaction of BAFF with its receptors.

Atomic view of the HIV-1 matrix lattice; implications on virus assembly and envelope incorporation

During the late phase of HIV type 1 (HIV-1) infection cycle, the virally encoded Gag polyproteins are targeted to the inner leaflet of the plasma membrane (PM) for assembly, formation of immature particles, and virus release. Gag binding to the PM is mediated by interactions of the N-terminally myristoylated matrix (myrMA) domain with phosphatidylinositol 4,5-bisphosphate. Formation of a myrMA lattice on the PM is an obligatory step for the assembly of immature HIV-1 particles and envelope (Env) incorporation. Atomic details of the myrMA lattice and how it mediates Env incorporation are lacking. Herein, we present the X-ray structure of myrMA at 2.15 Å. The myrMA lattice is arranged as a hexamer of trimers with a central hole, thought to accommodate the C-terminal tail of Env to promote incorporation into virions. The trimer–trimer interactions in the lattice are mediated by the N-terminal loop of one myrMA molecule and α-helices I–II, as well as the 310 helix of a myrMA molecule from an adjacent trimer. We provide evidence that substitution of MA residues Leu13 and Leu31, previously shown to have adverse effects on Env incorporation, induced a conformational change in myrMA, which may destabilize the trimer–trimer interactions within the lattice. We also show that PI(4,5)P2 is capable of binding to alternating sites on MA, consistent with an MA–membrane binding mechanism during assembly of the immature particle and upon maturation. Altogether, these findings advance our understanding of a key mechanism in HIV-1 particle assembly.

Cryo-EM structures of infectious bursal disease viruses with different virulences provide insights into their assembly and invasion

Infectious bursal disease virus (IBDV) causes a highly contagious immunosuppressive disease in chickens, resulting in significant economic losses. The very virulent IBDV strain (vvIBDV) causes high mortality and cannot adapt to cell culture. In contrast, attenuated strains of IBDV are nonpathogenic to chickens and can replicate in cell culture. Although the crystal structure of T = 1 subviral particles (SVP) has been reported, the structures of intact IBDV virions with different virulences remain elusive. Here, we determined the cryo-electron microscopy (cryo-EM) structures of the vvIBDV Gx strain and its attenuated IBDV strain Gt at resolutions of 3.3 Å and 3.2 Å, respectively. Compared with the structure of T = 1 SVP, IBDV contains several conserved structural elements unique to the T = 13 virion. Notably, the N-terminus of VP2, which is disordered in the SVP, interacts with the SF strand of VP2 from its neighboring trimer, completing the β-sheet of the S domain. This interaction helps to form a contact network by tethering the adjacent VP2 trimers and contributes to the assembly and stability of the IBDV virion. Structural comparison of the Gx and Gt strains indicates that H253 and T284 in the VP2 P domain of Gt, in contrast to Gx, form a hydrogen bond with a positively charged surface. This suggests that the combined mutations Q253H/A284T and the associated structural electrostatic features of the attenuated Gt strain may contribute to adaptation to cell culture. Furthermore, a negatively charged groove in VP2, containing an integrin binding IDA motif that is critical for virus attachment, was speculated to play a functional role in the entry of IBDV.

Reconstruction of the Fas-Based Death-Inducing Signaling Complex (DISC) Using a Protein–Protein Docking Meta-Approach

The death-inducing signaling complex (DISC) is a fundamental multiprotein complex, which triggers the extrinsic apoptosis pathway through stimulation by death ligands. DISC consists of different death domain (DD) and death effector domain (DED) containing proteins such as the death receptor Fas (CD95) in complex with FADD, procaspase-8, and cFLIP. Despite many experimental and theoretical studies in this area, there is no global agreement neither on the DISC architecture nor on the mechanism of action of the involved species. In the current work, we have tried to reconstruct the DISC structure by identifying key protein interactions using a new protein–protein docking meta-approach. We combined the benefits of five of the most employed protein–protein docking engines, HADDOCK, ClusPro, HDOCK, GRAMM-X, and ZDOCK, in order to improve the accuracy of the predicted docking complexes. Free energy of binding and hot spot interacting residues were calculated and determined for each protein–protein interaction using molecular mechanics generalized Born surface area and alanine scanning techniques, respectively. In addition, a series of in-cellulo protein-fragment complementation assays were conducted to validate the protein–protein docking procedure. The results show that the DISC formation initiates by dimerization of adjacent FasDD trimers followed by recruitment of FADD through homotypic DD interactions with the oligomerized death receptor. Furthermore, the in-silico outcomes indicate that cFLIP cannot bind directly to FADD; instead, cFLIP recruitment to the DISC is a hierarchical and cooperative process where FADD initially recruits procaspase-8, which in turn recruits and heterodimerizes with cFLIP. Finally, a possible structure of the entire DISC is proposed based on the docking results.

Augmenting the Immune Response against a Stabilized HIV-1 Clade C Envelope Trimer by Silica Nanoparticle Delivery.

The delivery of HIV-1 envelope (Env) trimer-based immunogens on the surface of nanoparticles holds promise to promote immunogenicity with the aim of inducing a potent, durable and broad neutralizing antibody (bnAb) response. Towards that goal, we examined the covalent conjugation of Env to 100 nm and 200 nm silica nanoparticles (SiNPs) to optimize conjugation density and attachment stability. Env was redesigned to enable site-specific cysteine-mediated covalent conjugation while maintaining its structural integrity and antigenicity. Env was anchored to different sized SiNPs with a calculated spacing of 15 nm between adjacent trimers. Both particle sizes exhibited high in vitro stability over a seven-day period. After attachment, 100 nm particles showed better colloidal stability compared to 200 nm particles. Importantly, the antigenic profile of Env was not impaired by surface attachment, indicating that the quaternary structure was maintained. In vitro Env uptake by dendritic cells was significantly enhanced when Env was delivered on the surface of nanoparticles compared to soluble Env. Furthermore, multivalent Env displayed efficiently activated B cells even at Env concentrations in the low nanomolar range. In mice, antibody responses to nanoparticle-coupled Env were stronger compared to the free protein and had equivalent effects at lower doses and without adjuvant.

Homochiral to heterochiral transition in a pentahelicene monolayer on Bi(111).

We report the nucleation and two dimensional (2D) crystallization of the helical aromatic hydrocarbon pentahelicene ([5]H) on the semimetallic Bi(111) surface studied via low-temperature scanning tunneling microscopy. Individual homochiral dimers and heterochiral trimers appear on the substrate at a low coverage. With an increase in the coverage, a chiral phase transition takes place from the 2D conglomerate of [5]H dimers to the 2D racemate of [5]H trimers. The heterochiral [5]H trimers reveal a wavy arrangement due to the swing of 5[H] trimer rows after every second or third trimers. The swing mechanism of the trimer rows can be attributed to the steric repulsion between the adjacent trimers with same handedness.

Structure and function of bacteriophage CBA120 ORF211 (TSP2), the determinant of phage specificity towards E. coli O157:H7

The genome of Escherichia coli O157:H7 bacteriophage vB_EcoM_CBA120 encodes four distinct tailspike proteins (TSPs). The four TSPs, TSP1-4, attach to the phage baseplate forming a branched structure. We report the 1.9 Å resolution crystal structure of TSP2 (ORF211), the TSP that confers phage specificity towards E. coli O157:H7. The structure shows that the N-terminal 168 residues involved in TSPs complex assembly are disordered in the absence of partner proteins. The ensuing head domain contains only the first of two fold modules seen in other phage vB_EcoM_CBA120 TSPs. The catalytic site resides in a cleft at the interface between adjacent trimer subunits, where Asp506, Glu568, and Asp571 are located in close proximity. Replacement of Asp506 and Asp571 for alanine residues abolishes enzyme activity, thus identifying the acid/base catalytic machinery. However, activity remains intact when Asp506 and Asp571 are mutated into asparagine residues. Analysis of additional site-directed mutants in the background of the D506N:D571N mutant suggests engagement of an alternative catalytic apparatus comprising Glu568 and Tyr623. Finally, we demonstrate the catalytic role of two interacting glutamate residues of TSP1, located in a cleft between two trimer subunits, Glu456 and Glu483, underscoring the diversity of the catalytic apparatus employed by phage vB_EcoM_CBA120 TSPs.

Two three-dimensional Sc(III)-MOFs: Synthesis, crystal structure and catalytic property

Two new three-dimensional (3D) Sc(III)-based metal organic frameworks (MOFs) [Sc2(OH)2(TDC)2]·4H2O (1) and [Sc3O(fum)3(H2O)3(NO3)]·DMF (2) (H2TDC = 2,5-thiophenedicarboxylic acid, H2fum = fumaric acid) have been hydrothermally synthesized. For 1, TDC2− ligands connect infinite one-dimensional (1D) -Sc-OH-Sc-OH- chains to form open-framework with rhombic channels along the a axis. In 2, flexible fum2− ligands link adjacent trimer [Sc3O(COO)6(H2O)3] secondary building units showing hexagonal-closed circular pores with a diameter of approximate 9.9 Å. Furthermore, the desolvated material la can retain the framework, presenting good catalytic activity, significant universality and substrate-selectivity for Strecker-type reactions of trimethylsilyl cyanide (TMSCN) and various aromatic aldimines, especially for N-benzyl 4-methoxybenzaldimine.

How small-molecule inhibitors of dengue-virus infection interfere with viral membrane fusion.

Dengue virus (DV) is a compact, icosahedrally symmetric, enveloped particle, covered by 90 dimers of envelope protein (E), which mediates viral attachment and membrane fusion. Fusion requires a dimer-to-trimer transition and membrane engagement of hydrophobic 'fusion loops'. We previously characterized the steps in membrane fusion for the related West Nile virus (WNV), using recombinant, WNV virus-like particles (VLPs) for single-particle experiments (Chao et al., 2014). Trimerization and membrane engagement are rate-limiting; fusion requires at least two adjacent trimers; availability of competent monomers within the contact zone between virus and target membrane creates a trimerization bottleneck. We now report an extension of that work to dengue VLPs, from all four serotypes, finding an essentially similar mechanism. Small-molecule inhibitors of dengue virus infection that target E block its fusion-inducing conformational change. We show that ~12-14 bound molecules per particle (~20-25% occupancy) completely prevent fusion, consistent with the proposed mechanism.

Cryospectroscopy Studies of Intact Light-Harvesting Antennas Reveal Empirical Electronic Energy Transitions in Two Cyanobacteria Species

Understanding of electronic energy transition (EET) mechanisms from the light-harvesting unit to the reaction center in a natural system has been limited by (a) the use of conventional transient time-resolved spectroscopy at room temperature, which result in high signal-to-noise ratio and (b) examining extracts instead of intact light-harvesting units. Here, we report previously unknown differences and new insight in EET of two cyanobacteria species, Acaryochloris marina and Thermosynechoccocus vulcanus, which have been found only after using UV-vis, hole-burning, and fluorescence spectroscopy at ultralow temperature and examining their intact light-harvesting unit, phycobilisomes (PBS). Although the exciton formation is similarly induced by photoexcitation of chromophore assemblies in phycocyanin (PC) and allophycocyanin (APC) in PBSs of both species, the EET mechanisms are totally different, being adiabatic in A. marina and nonadiabatic in T. vulcanus. The PBS of A. marina has only one APC trimer and energy transfer is through coupling of α84 in APC with β84 in adjacent PC. In T. vulcanus, the PBS has three components: coupling between APC core and the entire PC rod and couplings of β-β18 and of LCM to β in the adjacent APC-like trimer. A total of 80% of the excitation energy is trapped in the coupling β-β18 and regulates the flow of energy from the high- to low-level terminal electronic transition emitter β-LCM. All these details cannot be observed at room temperature and in extracted units.

Distinct Mechanism for the Formation of the Ribonucleoprotein Complex of Tomato Spotted Wilt Virus.

The Tomato spotted wilt virus (TSWV) belongs to the Tospovirus genus of the Bunyaviridae family and represents the sole plant-infecting group within bunyavirus. TSWV encodes a nucleocapsid protein (N) which encapsidates the RNA genome to form a ribonucleoprotein complex (RNP). In addition, the N has multiple roles during the infection of plant cells. Here, we report the crystal structure of the full-length TSWV N. The N features a body domain consisting of an N-lobe and a C-lobe. These lobes clamp a positively charged groove which may constitute the RNA binding site. Furthermore, the body domains are flanked by N- and C-terminal arms which mediate homotypic interactions to the neighboring subunits, resulting in a ring-shaped N trimer. Interestingly, the C terminus of one protomer forms an additional interaction with the protomer of an adjacent trimer in the crystal, which may constitute a higher-order oligomerization contact. In this way, this study provides insights into the structure and trimeric assembly of TSWV N, which help to explain previous functional findings, but also suggests distinct N interactions within a higher-order RNP.IMPORTANCE TSWV is one of the most devastating plant pathogens that cause severe diseases in numerous agronomic and ornamental crops worldwide. TSWV is also the prototypic member of the Tospovirus genus, which is the sole group of plant-infecting viruses in the bunyavirus family. This study determined the structure of full-length TSWV N in an oligomeric state. The structural observations explain previously identified biological properties of TSWV N. Most importantly, the additional homotypic interaction between the C terminus of one protomer with another protomer indicates that there is a distinct mechanism of RNP formation in the bunyavirus family, thereby enhancing the current knowledge of negative-sense single-stranded RNA virus-encoded N. TSWV N is the last remaining representative N with an unknown structure in the bunyavirus family. Combined with previous studies, the structure of TSWV N helps to build a complete picture of the bunyavirus-encoded N family and reveals a close evolutionary relationship between orthobunyavirus, phlebovirus, hantavirus, and tospovirus.

Adsorption and dynamics of Si atoms at the monolayer Pb/Si(111) surface

In this work, we studied the adsorption behavior of deposited Si atoms along with their diffusion and other dynamic processes on a Pb monolayer-covered Si(111) surface from 125-230 K using a variable-temperature scanning tunneling microscope (STM). The Pb-covered Si(111) surface form a low-symmetry row-like structure in this temperature range and the Si atoms bind favorably to two specific on-top sites (T1A and T1B) on the trimer row after deposition at the sample temperature of 125 K. The Si atoms were immobile at low temperatures and started to switch between the two neighboring T1A and T1B sites within the same trimer when the temperature was raised to 150 K. When the temperature was raised above 160 K, the adsorbed Si atoms could hop to other trimers along the same trimer row. Below 170 K, short hops to adjacent trimers dominated, but long hops dominated at temperatures above 170 K. The activation energy and prefactor for the Si atoms diffusion were derived through analysis of continuous-time imaging at temperatures from 160-174 K. In addition, irreversible aggregation of single Si atoms into Si clusters started to occur at the phase boundaries or defective sites at temperatures above 170 K. This study provides crucial information for understanding the very initial stage of nucleation and growth behavior of epitaxial Si layers on a Pb-covered Si(111) surface. In addition, our study provides strong evidence for breaking in the mirror symmetry in the Pb-covered structure, which has implication for the atomic model of this controversial structure.

Single Particle Kinetic and Electron Cryo-Microscopy Studies of Flavivirus Membrane Fusion

Flaviviruses (a family that includes West Nile, Dengue, Tick-borne encephalitis and Zika viruses) use the envelope protein E, to promote membrane fusion during viral entry by undergoing a low-pH triggered conformational reorganization. We have examined the mechanism of WNV fusion and sought insight for intermediates during the conformational transition by following hemifusion of WNV virus-like particles in a single particle format, with small molecule inhibitors and using single particle electron cryo-microscopy (Cryo-EM). At the level of individual E subunits, trimer formation and membrane engagement of three fold clustered fusion loops is a kinetic bottleneck. Simulation of the kinetics indicates hemifusion requires at least two adjacent trimers, and that the availability of competent monomers within the contact zone between virus and target membrane is limiting. We present an initial Cryo-EM reconstruction of the full length post-fusion trimer and discuss how these visualization of the transmembrane regions, completed by further kinetic studies can describe principles of flavivirus entry generally applicable to mechanisms of membrane fusion.

Architecture of the Yeast Mitochondrial Iron-Sulfur Cluster Assembly Machinery: THE SUB-COMPLEX FORMED BY THE IRON DONOR, Yfh1 PROTEIN, AND THE SCAFFOLD, Isu1 PROTEIN

The biosynthesis of Fe-S clusters is a vital process involving the delivery of elemental iron and sulfur to scaffold proteins via molecular interactions that are still poorly defined. We reconstituted a stable, functional complex consisting of the iron donor, Yfh1 (yeast frataxin homologue 1), and the Fe-S cluster scaffold, Isu1, with 1:1 stoichiometry, [Yfh1]24·[Isu1]24 Using negative staining transmission EM and single particle analysis, we obtained a three-dimensional reconstruction of this complex at a resolution of ∼17 Å. In addition, via chemical cross-linking, limited proteolysis, and mass spectrometry, we identified protein-protein interaction surfaces within the complex. The data together reveal that [Yfh1]24·[Isu1]24 is a roughly cubic macromolecule consisting of one symmetric Isu1 trimer binding on top of one symmetric Yfh1 trimer at each of its eight vertices. Furthermore, molecular modeling suggests that two subunits of the cysteine desulfurase, Nfs1, may bind symmetrically on top of two adjacent Isu1 trimers in a manner that creates two putative [2Fe-2S] cluster assembly centers. In each center, conserved amino acids known to be involved in sulfur and iron donation by Nfs1 and Yfh1, respectively, are in close proximity to the Fe-S cluster-coordinating residues of Isu1. We suggest that this architecture is suitable to ensure concerted and protected transfer of potentially toxic iron and sulfur atoms to Isu1 during Fe-S cluster assembly.

Sequential conformational rearrangements in flavivirus membrane fusion.

The West Nile Virus (WNV) envelope protein, E, promotes membrane fusion during viral cell entry by undergoing a low-pH triggered conformational reorganization. We have examined the mechanism of WNV fusion and sought evidence for potential intermediates during the conformational transition by following hemifusion of WNV virus-like particles (VLPs) in a single particle format. We have introduced specific mutations into E, to relate their influence on fusion kinetics to structural features of the protein. At the level of individual E subunits, trimer formation and membrane engagement of the threefold clustered fusion loops are rate-limiting. Hemifusion requires at least two adjacent trimers. Simulation of the kinetics indicates that availability of competent monomers within the contact zone between virus and target membrane makes trimerization a bottleneck in hemifusion. We discuss the implications of the model we have derived for mechanisms of membrane fusion in other contexts.

A novel packing arrangement of AcrB in the lipid bilayer membrane

The central component AcrB of the Escherichia coli drug efflux complex AcrA–AcrB–TolC has been extensively investigated by X-ray crystallography of detergent–protein 3-D crystals. In these crystals, AcrB packs as trimers – the functional unit. We visualized the AcrB–AcrB interaction in its native environment by examining E. coli lipid reconstituted 2-D crystals, which were overwhelmingly formed by asymmetric trimers stabilized by strongly-interacting monomers from adjacent trimers. Most interestingly, we observed lattices formed by an arrangement of AcrB monomers distinct from that in traditional trimers. This hitherto unobserved packing, might play a role in the biogenesis of trimeric AcrB.

Clarifying and illustrating the electronic energy transfer pathways in trimeric and hexameric aggregation state of cyanobacteria allophycocyanin within the framework of Förster theory.

Within the framework of the Förster theory, the electronic excitation energy transfer pathways in the cyanobacteria allophycocyanin (APC) trimer and hexamer were studied. The associated physical quantities (i.e., excitation energy, oscillator strength, and transition dipole moments) of the phycocyanobilins (PCBs) located in APC were calculated at time-dependent density functional theory (TDDFT) level of theory. To estimate the influence of protein environment on the preceding calculated physical quantities, the long-range interactions were approximately considered with the polarizable continuum model at the TDDFT level of theory, and the short-range interaction caused by surrounding aspartate residue of PCBs were taken into account as well. The shortest energy transfer time calculated in the framework of the Förster model at TDDFT/B3LYP/6-31+G* level of theory are about 0.10 ps in the APC trimer and about 170 ps in the APC monomer, which are in qualitative agreement with the experimental finding that a very fast lifetime of 0.43-0.44 ps in APC trimers, whereas its monomers lacked any corresponding lifetime. These results suggest that the lifetime of 0.43-0.44 ps in the APC trimers determined by Sharkov et al. was most likely attributed to the energy transfer of α(1) -84 ↔ β(3) -84 (0.23 ps), β(1) -84 ↔ α(2) -84 (0.11 ps) or β(2) -84 ↔ α(3) -84 (0.10 ps). So far, no experimental or theoretical energy transfer rates between two APC trimmers were reported, our calculations predict that the predominate energy transfer pathway between APC trimers is likely to occur from α(3) -84 in one trimer to α(5) -84 in an adjacent trimer with a rate of 32.51 ps.

Why are the 2-oxoacid dehydrogenase complexes so large? Generation of an active trimeric complex.

The four-component polypeptides of the 2-oxoacid dehydrogenase complex from the thermophilic archaeon Thermoplasma acidophilum assemble to give an active multienzyme complex possessing activity with the branched-chain 2-oxoacids derived from leucine, isoleucine and valine, and with pyruvate. The dihydrolipoyl acyl-transferase (E2) core of the complex is composed of identical trimer-forming units that assemble into a novel 42-mer structure comprising octahedral and icosahedral geometric aspects. From our previously determined structure of this catalytic core, the inter-trimer interactions involve a tyrosine residue near the C-terminus secured in a hydrophobic pocket of an adjacent trimer like a ball-and-socket joint. In the present study, we have deleted the five C-terminal amino acids of the E2 polypeptide (IIYEI) and shown by equilibrium centrifugation that it now only assembles into a trimeric enzyme. This was confirmed by SAXS analysis, although this technique showed the presence of approximately 20% hexamers. The crystal structure of the trimeric truncated E2 core has been determined and shown to be virtually identical with the ones observed in the 42-mer, demonstrating that removal of the C-terminal anchor does not significantly affect the individual monomer or trimer structures. The truncated E2 is still able to bind both 2-oxoacid decarboxylase (E1) and dihydrolipoamide dehydrogenase (E3) components to give an active complex with catalytic activity similar to the native multienzyme complex. This is the first report of an active mini-complex for this enzyme, and raises the question of why all 2-oxoacid dehydrogenase complexes assemble into such large structures.

Stochastic Fusion Simulations and Experiments Suggest Passive and Active Roles of Hemagglutinin during Membrane Fusion

Influenza enters the host cell cytoplasm by fusing the viral and host membrane together. Fusion is mediated by hemagglutinin (HA) trimers that undergo conformational change when acidified in the endosome. It is currently debated how many HA trimers, w, and how many conformationally changed HA trimers, q, are minimally required for fusion. Conclusions vary because there are three common approaches for determining w and q from fusion data. One approach correlates the fusion rate with the fraction of fusogenic HA trimers and leads to the conclusion that one HA trimer is required for fusion. A second approach correlates the fusion rate with the total concentration of fusogenic HA trimers and indicates that more than one HA trimer is required. A third approach applies statistical models to fusion rate data obtained at a single HA density to establish w or q and suggests that more than one HA trimer is required. In this work, all three approaches are investigated through stochastic fusion simulations and experiments to elucidate the roles of HA and its ability to bend the target membrane during fusion. We find that the apparent discrepancies among the results from the various approaches may be resolved if nonfusogenic HA participates in fusion through interactions with a fusogenic HA. Our results, based on H3 and H1 serotypes, suggest that three adjacent HA trimers and one conformationally changed HA trimer are minimally required to induce membrane fusion (w = 3 and q = 1).

The structure of 3-(diethylborylethynyl)pyridine: a nonplanarly arranged cyclic trimer.

3-(Diethylborylethynyl)pyridines 2 assemble into a cyclic trimer stabilized via intermolecular boron–nitrogen coordination bonds both in solution and in the crystalline state. The outstanding structural features of the methoxy derivative 2b in the crystalline state are that (1) two pyridine rings (P1 and P2) of the cyclic trimer of 2b are almost coplanar, and the third pyridine ring (P3) is largely bent away from P1 and P2, and (2) P3 of the cyclic trimer stacks in a face-to-face fashion with one of the pyridine rings (P3’) of an adjacent cyclic trimer. The crystallographic study revealed that the conformation of the cyclic trimer is flexible enough to be affected by the crystal packing.