Structures of foot-and-mouth disease virus with bovine neutralizing antibodies reveal the determinant of intra-serotype cross-neutralization

Foot-and-mouth disease virus (FMDV) exhibits broad antigenic diversity with poor intra-serotype cross-neutralizing activity. Studies of the determinant involved in this diversity are essential for the development of broadly protective vaccines. In this work, we isolated a bovine antibody, designated R55, that displays cross-reaction with both FMDV A/AF/72 (hereafter named FMDV-AAF) and FMDV A/WH/09 (hereafter named FMDV-AWH) but only has a neutralizing effect on FMDV-AWH. Near-atomic resolution structures of FMDV-AAF-R55 and FMDV-AWH-R55 show that R55 engages the capsids of both FMDV-AAF and FMDV-AWH near the icosahedral threefold axis and binds to the βB and BC/HI-loops of VP2 and to the B-B knob of VP3. The common interaction residues are highly conserved, which is the major determinant for cross-reaction with both FMDV-AAF and FMDV-AWH. In addition, the cryo-EM structure of the FMDV-AWH-R55 complex also shows that R55 binds to VP3 E70 located at the VP3 BC-loop in an adjacent pentamer, which enhances the acid and thermal stabilities of the viral capsid. This may prevent capsid dissociation and genome release into host cells, eventually leading to neutralization of the viral infection. In contrast, R55 binds only to the FMDV-AAF capsid within one pentamer due to the VP3 E70G variation, which neither enhances capsid stability nor neutralizes FMDV-AAF infection. The VP3 E70G mutation is the major determinant involved in the neutralizing differences between FMDV-AWH and FMDV-AAF. The crucial amino acid VP3 E70 is a key component of the neutralizing epitopes, which may aid in the development of broadly protective vaccines. Importance Foot-and-mouth disease virus (FMDV) causes a highly contagious and economically devastating disease in cloven-hoofed animals, and neutralizing antibodies play critical roles in the defense against viral infections. Here, we isolated a bovine antibody (R55) using the single B cell antibody isolation technique. Enzyme-linked immunosorbent assays (ELISA) and virus neutralization tests (VNT) showed that R55 displays cross-reactions with both FMDV-AWH and FMDV-AAF but only has a neutralizing effect on FMDV-AWH. Cryo-EM structures, fluorescence-based thermal stability assays and acid stability assays showed that R55 engages the capsid of FMDV-AWH near the icosahedral threefold axis and informs an interpentamer epitope, which overstabilizes virions to hinder capsid dissociation to release the genome, eventually leading to neutralization of viral infection. The crucial amino acid VP3 E70 forms a key component of neutralizing epitopes, and the determination of the VP3 E70G mutation involved in the neutralizing differences between FMDV-AWH and FMDV-AAF could aid in the development of broadly protective vaccines.

Synthesis and structure determination of racemic (Δ/Λ)-tris(ethylenediamine)cobalt(III) trichloride hemi(hexaaquasodium chloride)

The synthesis and crystal structure of the title racemic compound, [Co(C2H8N2)3]Cl3.{[Na(H2O)6]Cl}0.5, are reported. The trivalent cobalt atom, which resides on a crystallographic threefold axis, is chelated by a single ethylene diamine (en) ligand and yields the tris-chelate [Co(en)3]3+ cation with distorted octahedral geometry after the application of crystal symmetry. The sodium cation (site symmetry \overline{3}), has a single water molecule bound to it in the asymmetric unit and yields a distorted, octahedrally coordinated hydrated [Na(H2O)6]+ cation after the application of symmetry. One of the chloride ions lies on a general position and the other has \overline{3} site symmetry. An extensive array of C—H...O, N—H...Cl and O—H...Cl hydrogen bonds exists between the ethylene diamine ligands, the water molecules of hydration, and the anions present, thereby furnishing solid-state stability.

Cryo-EM structure of a thermostable bacterial nanocompartment

Protein nanocompartments are widespread in bacteria and archaea, but their functions are not yet well understood. Here, the cryo-EM structure of a nanocompartment from the thermophilic bacterium Thermotoga maritima is reported at 2.0 Å resolution. The high resolution of this structure shows that interactions in the E-loop domain may be important for the thermostability of the nanocompartment assembly. Also, the channels at the fivefold axis, threefold axis and dimer interface are assessed for their ability to transport iron. Finally, an unexpected flavin ligand was identified on the exterior of the shell, indicating that this nanocompartment may also play a direct role in iron metabolism.

Hydrogen-bonding in mono-, di- and tetramethylammonium dihydrogenphosphites

The crystal structures of methylammonium and dimethylammonium dihydrogenphosphite (MA⋅H2PO3, I2/a and DMA⋅H2PO3, P 2 1 / c $P{2}_{1}/c$ ) are built of infinite chains of hydrogen bonded H 2 P O 3 − ${\mathrm{H}}_{\mathrm{2}}\mathrm{P}{\mathrm{O}}_{\mathrm{3}}^{-}$ anions. The chains are connected by the ammonium cations via hydrogen bonding to di- (DMA⋅H2PO3) and triperiodic (MA⋅H2PO3) networks. Tetramethylammonium dihydrogenphosphite monohydrate (TMA⋅H2PO3⋅H2O) features temperature dependent dimorphism. The crystal structure of the high-temperature (HT, cubic P213) and low-temperature (LT, orthorhombic P212121) phases were determined at 150 and 100 K, respectively. The hydrogen bonding network in the HT phase is disordered, with H 2 P O 3 − ${\mathrm{H}}_{\mathrm{2}}\mathrm{P}{\mathrm{O}}_{\mathrm{3}}^{-}$ and H2O being located on a threefold axis and is ordered in the LT phase. On cooling, the point symmetry is reduced by an index of 3. The lost symmetry is retained as twin operations, leading to threefold twinning by pseudo-merohedry. The hydrogen-bonding networks of the HT and LT phases can be represented by undirected and directed quotient graphs, respectively.

Designed ferritin nanocages displaying trimeric TRAIL and tumor-targeting peptides confer superior anti-tumor efficacy

TRAIL is considered a promising target for cancer therapy because it mediates activation of the extrinsic apoptosis pathway in a tumor-specific manner by binding to and trimerizing its functional receptors, DR4 or DR5. Although recombinant human TRAIL has shown high potency and specificity for killing cancer cells in preclinical studies, it has failed in multiple clinical trials for several reasons, including a very short half-life mainly caused by instability of the monomeric form of TRAIL and rapid renal clearance of the off-targeted TRAIL. To overcome such obstacles, we developed a TRAIL-active trimer nanocage (TRAIL-ATNC) that presents the TRAIL ligand in its trimer-like conformation by connecting it to a triple helix sequence that links to the threefold axis of the ferritin nanocage. We also ligated the tumor-targeting peptide, IL4rP, to TRAIL-ATNC to enhance tumor targeting. The developed TRAIL-ATNCIL4rP showed enhanced agonistic activity compared with monomeric TRAIL. The in vivo serum half-life of TRAIL-ATNCIL4rP was ~ 16-times longer than that of native TRAIL. As a consequence of these properties, TRAIL-ATNCIL4rP exhibited efficacy as an anti-tumor agent in vivo against xenograft breast cancer as well as orthotopic pancreatic cancer models, highlighting the promise of this system for development as novel therapeutics against cancer.

The Crystal Structures of Thermomyces (Humicola) Lanuginosa Lipase in Complex with Enzymatic Reactants

Aim: To understand the details of the action of fungal lipase and the mechanism for its observed interfacial activation. Background: Fungal lipase, crucial to biotechnology, functions at the lipid - water interface where it undergoes a poorly understood interfacial activation. Biochemical factors influencing its activation and inhibition are also poorly understood. This study provides a basis for its activity and a plausible mechanism for interfacial activation. Objective: To determine the structures of fungal lipase in different crystal forms in complex with their enzymatic reactants and inhibitors. Method: X-ray crystallography. Results: Thermomyces lanuginosa lipase was visualized in three crystal forms, of space groups H32, P21 and I222 at 1.3 to 1.45 Å resolution. Rhombohedral crystals have one molecule, lacking segment 241 to 252, as an asymmetric unit, with molecules organized as two trimers. Monoclinic crystals’ asymmetric unit is six intact molecules organized as two, nearly identical trimers, each exhibiting an NCS threefold axis. The “lid” helix was consistently closed. Oligomerization into trimers creates an internal hydrophobic cavity where catalysis occurs. In monoclinic and orthorhombic crystals, active site serines were esterified to fatty acids. Lipase had bound within their trimeric, hydrophobic cavities 1,3-diacylglycerols with fatty acid chain lengths of about 18 carbons. Conclusions: Results suggest trimers are likely the active form of the enzyme at the lipid-water interface. Formation of trimers may provide an explanation for “interfacial activation”.

Crystal structure of Ag-exchanged levyne intergrown with erionite: Single-crystal X-ray diffraction and Molecular Dynamics simulations

The modification of natural zeolites via ion exchange is an efficient technique used to improve their performances and tune their properties for specific applications. In this study, a natural levyne-Ca intergrown with erionite was fully exchanged by Ag+ and its structure [with idealized chemical composition Ag6(Si,Al)18O36·18H2O] was investigated by combining a theoretical and experimental approach. Single-crystal X-ray diffraction data demonstrated that Ag-levyne maintained the R3m space group, characteristic of the natural levyne. Ag ions distribute over partially occupied sites along the threefold axis and, differently from the pristine material, at the wall of the 8-membered ring window of the lev cavity. The lack of ~30% of Ag ions that could not be located by the structural refinement is ascribed to the strong disorder of the extraframework occupants. The structural results obtained by Molecular Dynamics simulations are in overall agreement with the experimental data and showed that, on average, Ag+ is surrounded by ~2 H2O and 1 framework oxygen at distances between 2.43 and 2.6 Å. Molecular Dynamics trajectories indicate that the occurrence of silver inside the D6R cage depends on the water content: silver occupancy of D6R cages is estimated to be 83, 30, and 0% when the structure contains 3, 2.5, and 2 H2O per Ag ion, respectively. The cation-exchange process, as demonstrated by scanning electron microscopy and energy-dispersive spectroscopy (SEM-EDS) spectrometry, affects the intergrown erionite as well. A structural characterization of the Ag-erionite phase (with dimension <100 μm) was possible by means of a CuKα micro-focus source: structure solution pointed to P63/mmc space group, indicating no change with respect to natural erionite. In agreement with previous studies, K ions in the cancrinite cage could not be exchanged, whereas Ag+ is found in the eri cavity.

4-[(1E)-({[(Benzylsulfanyl)methanethioyl]amino}imino)methyl]benzene-1,3-diol chloroform hemisolvate: crystal structure, Hirshfeld surface analysis and computational study

The title hydrazine carbodithioate chloroform hemisolvate, 2C15H14N2O2S2·CHCl3, comprises two independent hydrazine carbodithioate molecules, A and B, and a chloroform molecule; the latter is statistically disordered about its molecular threefold axis. The common features of the organic molecules include an almost planar, central CN2S2 chromophore [r.m.s. deviation = 0.0203 Å (A) and 0.0080 Å (B)], an E configuration about the imine bond and an intramolecular hydroxyl-O—H...N(imine) hydrogen bond. The major conformational difference between the molecules is seen in the relative dispositions of the phenyl rings as indicated by the values of the dihedral angles between the central plane and phenyl ring of 71.21 (6)° (A) and 54.73 (7)° (B). Finally, a difference is seen in the disposition of the outer hydroxyl-H atoms, having opposite relative orientations. In the calculated gas-phase structure, the entire molecule is planar with the exception of the perpendicular phenyl ring. In the molecular packing, the A and B molecules assemble into a two-molecule aggregate via N—H...S hydrogen bonds and eight-membered {...HNCS}2 synthons. The dimeric assemblies are connected into supramolecular chains via hydroxyl-O—H...O(hydroxyl) hydrogen bonds and these are linked into a double-chain through hydroxy-O—H...π(phenyl) interactions. The double-chains are connected into a three-dimensional architecture through phenyl-C—H...O(hydroxyl) and phenyl-C—H...π(phenyl) interactions. The overall assembly defines columns along the a-axis direction in which reside the chloroform molecules, which are stabilized by chloroform–methine-C—H...S(thione) and phenyl-C—H...Cl contacts. The analysis of the calculated Hirshfeld surfaces, non-covalent interaction plots and interaction energies confirm the importance of the above-mentioned interactions, but also of cooperative, non-standard interactions such as π(benzene)...π(hydrogen-bond-mediated-ring) contacts.

Low-Temperature Ordering in the Cluster Compound (Bi8)Tl[AlCl4]3

The reaction of Bi, BiCl3, and TlCl in the ionic liquid [BMIm]Cl·4AlCl3 (BMIm = 1-n-butyl-3-methylimidazolium) at 180 °C yielded air-sensitive black crystals of (Bi8)Tl[AlCl4]3. X-ray diffraction on single crystals at room temperature revealed a structure containing [ Tl ( AlCl 4 ) 3 ] ∞ 1 2 − strands separated by isolated Bi82+ square antiprisms. The thallium(I) ion is coordinated by twelve Cl− ions of six [AlCl4]− groups, resulting in a chain of face-sharing [TlCl12]11− icosahedra. The Bi82+ polycation is disordered, simulating a threefold axis through its center and overall hexagonal symmetry (space group P63/m). Slowly cooling the crystals to 170 K resulted in increased order in the Bi8 cluster orientations. An ordered structure model in a supercell with a’ = 2a, b’ = 2b, c’ = 3c and the space group P65 was refined. The structure resembles a hexagonal perovskite, with complex groups in place of simple ions.

Structural basis for activity of TRIC counter-ion channels in calcium release

Trimeric intracellular cation (TRIC) channels are thought to provide counter-ion currents that facilitate the active release of Ca2+from intracellular stores. TRIC activity is controlled by voltage and Ca2+modulation, but underlying mechanisms have remained unknown. Here we describe high-resolution crystal structures of vertebrate TRIC-A and TRIC-B channels, both in Ca2+-bound and Ca2+-free states, and we analyze conductance properties in structure-inspired mutagenesis experiments. The TRIC channels are symmetric trimers, wherein we find a pore in each protomer that is gated by a highly conserved lysine residue. In the resting state, Ca2+binding at the luminal surface of TRIC-A, on its threefold axis, stabilizes lysine blockage of the pores. During active Ca2+release, luminal Ca2+depletion removes inhibition to permit the lysine-bearing and voltage-sensing helix to move in response to consequent membrane hyperpolarization. Diacylglycerol is found at interprotomer interfaces, suggesting a role in metabolic control.