Pentameric Structure Research Articles

Supramolecularly assembled pentameric and octameric water clusters stabilized by a mixed complex of Ni(II)

Pyridine-2,4,6-tricarboxylic acid (ptcH 3) reacts with Ni(NO 3) 2 · 6H 2O and 2,2′-bipyridyl (bpy) in 2:2:1 molar ratio to form green crystals with the empirical formula, {[Ni(ptcH) 2][Ni(bpy)(H 2O) 4]6H 2O} ( 1). Water molecules present are associated to form discrete pentameric and octameric clusters which form an intricate array of H-bonding interactions with carboxylate O atoms, leading to a H-bonded 3D open framework structure. The water octamer takes the shape of a hexameric chair to which two water molecules are attached at one and four positions, while the pentameric structure is linear. Both these structures are unprecedented and their shapes are commensurate with the overall structure of the complex. Thermal analysis, X-ray powder diffraction and X-ray structural analysis have been used to characterize this compound.

Stoichiometry and Pharmacology of Two Human α4β2 Nicotinic Receptor Types

The alpha4beta2 nicotinic acetylcholine receptor (nAChR) is the most abundant nAChR subtype in the brain, where it forms the high-affinity binding site for nicotine. The alpha4beta2 nAChR belongs to a gene family of ligand-gated ion channels that also includes muscle nAChRs, GABAA receptors, and glycine receptors and that assembles into pentameric structures. alpha4 and beta2 nAChR subunits expressed heterologously in Xenopus laevis oocytes assemble into a mixture of high- and low-affinity functional receptors, giving rise to biphasic ACh concentration-response curves (Zwart and Vijverberg, 1998; Buisson and Bertrand, 2001; Houlihan et al., 2001). High- and low-affinity alpha4beta2 nAChRs differ significantly in their functional and pharmacological properties (Zwart and Vijverberg, 1998; Buisson and Bertrand, 2001; Houlihan et al., 2001; Nelson et al., 2003) and result from the assembly of alpha4 and beta2 subunits into two distinct stoichiometric arrangements: (alpha4)2(beta2)3(high-affinity subtype) and (alpha4)3(beta2)2 (low-affinity subtype) (Nelson et al., 2003). In this study we have examined the functional and pharmacological properties of high- and low-affinity alpha4beta2 receptors using two-electrode voltage clamp procedures on Xenopus oocytes transfected with high (1:10) or low (10:1) ratios of alpha4/beta2 cDNAs, which yield high (1:10)- or low (10:1)- affinity receptors with monophasic ACh concentration- response curves. Furthermore, to determine the stoichiometry of high- and low-affinity receptors expressed heterologously by Xenopus oocytes, we have determined the stoichiometry of high- and low-affinity alpha4beta2 receptors by mutating a highly conserved hydrophobic residue in the middle (position 9') of the pore-lining domain, which increases agonist potency in a manner that allows predictions on subunit composition (Cooper et al., 1991; Revah et al., 1991; Labarca et al., 1995; Boorman et al., 2000).

The VP1 protein of human enterovirus 71 self-associates via an interaction domain spanning amino acids 66–297

Enterovirus 71 (EV71) is a major etiological agent of hand, foot, and mouth disease (HFMD). Several recent outbreaks of HFMD in East Asia were associated with neurological complications and numerous deaths. In 2000, an outbreak in Singapore afflicted thousands of children, resulting in four fatal cases from whom EV71 was isolated. The virus possesses four structural proteins VP1, VP2, VP3, and VP4, each of which is involved in forming the pentameric icosahedral structure of the virus. Here we report that the full-length VP1 structural protein of EV71 is capable of self-association. Dimerization of VP1 was tested using the yeast two-hybrid system, fluorescence resonance energy transfer (FRET) analysis, in vitro coupled transcription-translation binding assays, and mammalian cell transcription-translation experiments. Dimerization of various truncated versions of the VP1 protein was also studied by mutational analysis. Systematic deletions of parts of VP1 revealed that the region spanning amino acids 66-132 of VP1 contains the major dimerization domain. However, the region between amino acids 132 and 297 was indispensable, and contributed largely to increasing the strength of the interaction. This ability of EV71 VP1 to self-associate and to participate significantly in forming the characteristic icosahedral capsid strongly enhances the pathogenicity and stability of the virus to withstand the environment of the gastrointestinal tract.

Interactions of C-reactive protein with low-density lipoproteins: Implications for an active role of modified C-reactive protein in atherosclerosis

The interaction of C-reactive protein with low-density lipoprotein is considered to be one of the key properties that link C-reactive protein with atherosclerosis. However the data obtained to date are controversial, and hence make it difficult to conclude actual physiological or pathological impact of such interaction. The incompatible findings could be ascribed to the different structural state of C-reactive protein and/or low-density lipoprotein. We investigated in detail the interaction of various C-reactive protein isoforms with native and modified low-density lipoprotein. Our data showed “C-reactive protein” could indeed interact with each of native low-density lipoprotein, oxidized or enzymatically modified low-density lipoprotein, but that interaction occurs primarily when C-reactive protein is conformed in a modified form and not pentameric structure. Low level of modified C-reactive protein “contaminant” could confer C-reactive protein obvious low-density lipoprotein binding capacity. Interaction of modified C-reactive protein and low-density lipoprotein was mediated synergistically by both electrostatic association with ApoB and hydrophobic insertion into lipid layer. When complexed with modified C-reactive protein, macrophage binding/uptake of native and oxidized low-density lipoprotein was either increased 150% or decreased 35%, respectively. Thus the interaction of modified C-reactive protein with low-density lipoprotein may contribute to the regulation of low-density lipoprotein metabolism and foam cell formation in arterial wall. These results highlight an active role of modified C-reactive protein in atherosclerotic process.

First joint Italian–German purine club meeting “Progress in Purinergic Receptor Pharmacology and Function”

First joint Italian–German purine club meeting “Progress in Purinergic Receptor Pharmacology and Function”

A Flexible Approach to the Calculation of Resonance Energy Transfer Efficiency between Multiple Donors and Acceptors in Complex Geometries

Resonance energy transfer provides a practical way to measure distances in the range of 10–100 Å between sites in biological molecules. Although the relationship between the efficiency of energy transfer and the distance between sites is well described for a single pair of fluorophores, the situation is more difficult when more than two fluorophores are present. Using a Monte Carlo calculation scheme, we demonstrate how resonance energy transfer can be used to measure distances between fluorophores in complex geometries. We demonstrate the versatility of the approach by calculating the efficiency of energy transfer for individual fluorophores randomly distributed in two and three dimensions, for linked pairs of donors and acceptors and pentameric structures of five linked fluorophores. This approach can be used to relate the efficiency of energy transfer to the distances between fluorophores, R 0, molecular concentrations, laser power, and donor/acceptor ratios in ensembles of molecules or when many fluorophores are attached to a single molecule such as in multimeric proteins.

Synthesis and assembly of an adjuvanted Porphyromonas gingivalis fimbrial antigen fusion protein in plants

The gram-negative anaerobic oral bacterium Porphyromonas gingivalis initiates periodontal disease by binding to saliva-coated oral surfaces. To assess whether edible plants can synthesize biologically active P. gingivalis fimbrial antigen, for application as an oral vaccine, a cDNA fragment encoding the C-terminal binding portion of P. gingivalis fimbrial protein (FimA), was cloned into a plant expression vector immediately downstream of a cDNA fragment encoding the cholera toxin B subunit (CTB). The chimeric plasmid was transferred into potato ( Solanum tuberosum) cells and the ctb– fimA cDNA fragment detected in transformed leaf genomic DNA by PCR amplification methods. A novel protein band of 21 kDa was detected in transformed potato tuber extracts by immunoblot analysis. Oligomeric CTB–FimA (266–337) fusion protein was identified in the extracts through the binding of anti-CTX and anti-native fimbriae antibodies. The pentameric structure of CTB–FimA fusion protein was confirmed by ELISA measurements of G M1 ganglioside receptor binding. Quantification of the CTB–FimA fusion protein by ELISA indicated that the chimeric protein made up about 0.33% of total soluble tuber protein. The biosynthesis of immunologically detectable CTB–FimA fusion proteins and the assembly of fusion protein monomers into biologically active pentamers in transformed potato tuber tissues demonstrate the feasibility of synthesizing adjuvanted fimbrial protein in edible plants for development of adjuvanted mucosal vaccines against P. gingivalis generated periodontal disease.

Loss of Pentameric Symmetry in C-Reactive Protein Induces Interleukin-8 Secretion Through Peroxynitrite Signaling in Human Neutrophils

Plasma levels of C-reactive protein (CRP), nitrotyrosine, and interleukin-8 (IL-8) are known predictors of acute cardiovascular events. Peroxynitrite (ONOO-) may function as an intracellular signal for the production of IL-8; however, it is not known whether CRP regulates these events. Emerging evidence suggests that some bioactivities of CRP are expressed only when the pentameric structure of CRP is lost, resulting in formation of monomeric or modified CRP (mCRP). We studied the impact of human native CRP and bioengineered mCRP that cannot rearrange into the pentameric structure on ONOO- formation and ONOO--mediated IL-8 gene expression in human leukocytes. Incubation of human whole blood or isolated neutrophils with mCRP (0.1 to 100 microg/mL) for 4 hours increased IL-8 gene expression and secretion that was blocked approximately 70% by the NO synthase inhibitor Nomega-nitro-L-arginine methyl ester (L-NAME). In neutrophils, mCRP simultaneously increased superoxide production and endothelial nitric oxide synthase-mediated NO formation, leading to enhanced ONOO- formation, and consequently activation of nuclear factor-kappaB and activator protein-1. Native CRP had no detectable effect at 4 hours, whereas it enhanced IL-8 release after a 24-hour incubation that was blocked by L-NAME. An anti-CD16 antibody, but not an anti-CD32 antibody, produced 60% to 70% reductions in mCRP-stimulated NO formation and IL-8 release (both P<0.05). These results suggest that loss of the pentameric symmetry in CRP, resulting in formation of mCRP, leads to IL-8 release from human neutrophils via peroxynitrite-mediated activation of nuclear factor-kappaB and activator protein-1.

Structural, Potential Surface and Vibrational Spectroscopy Studies of Hypophosphorous Acid in the Gas Phase and Chain Conformation. A Theoretical Study

The potential energy surfaces (PES), molecular and vibrational structure of hypophosphours acid (HPA) were investigated by HF, MP2 and DFT-B3LYP level of theory using 6-31G** basis set. In order to approach solid state spectra we optimized mono-, di-, tri-, tetraand penta-mer structures and computed frequencies and intensities of the vibrational modes of VCD and IR. It is found that by increasing the number of HPA molecules in the chain, some of calculated vibrational frequencies approach to the experimental solid state values. The potential energy surfaces of HPA are calculated in a wide range on the plane perpendicular to the P-O bond where the hydrogen can rotate 360 degree through O=P-O-H dihedral angle. A circular valley is found going up and down on the plane, the valley is centered to the continuation of P-O bond. There are two minima at angles ±42.5 to the O=P-O plane giving two mirror conformers and one saddle point in between with a height of 284.87 cm at the angles 0 and a complex barrier with a height of 2089.40 cm at the angle 180 to the same plane. On top of the complex barrier, there is a small well with depth of 15.65 cm. To study the tunneling effect and pathway between the two conformers, the molecule is considered to have C S symmetry and a symmetric double minimum potential energy well with a barrier of 284.87 cm height in the middle. With the constructed potential, the torsional motion of P-O-H (Hindered Rotation) of the monomer, using variation method and harmonic oscillator wave functions is studied and IR frequencies and relative intensities of vibrational modes are calculated. Due to the width of the well and the trigonometric shape of the barrier, tunneling can occur at the ground state

Analysis of the stability and function of nucleoplasmin through cysteine mutants

Xenopus laevis nucleoplasmin is a pentameric nuclear chaperone. The relation between the structure and the multifunctional aspects of the molecule has not yet been clearly established. In the course of analysing a C-terminally His-tagged recombinant version of the region equivalent to the trypsin resistant core (r-NP142) of the molecule, we found that this domain exhibited a substantially decreased oligomerization potential. To better understand the role of the three cysteines of nucleoplasmin on its pentameric functional structure, we have selectively mutated these residues to serine and generated three mutants (C15S, C35S, and C45S) both for the complete recombinant nucleoplasmin (r-NP) and the truncated r-NP142 non-tagged forms. We demonstrate that there are no disulphide bridges stabilizing either the monomer or the pentamer. Neither C15S nor C35S has any structural effects, while the mutation C45S abolishes the ability of r-NP142 to pentamerize. This structural impairment suggests that hydrophobic interactions of Cys 45 are critical for the stability of the protein. Our studies allow to analyse for the first time the structural and functional properties of nucleoplasmin in its monomeric form.

A chimera encoding the fusion of an acetylcholine-binding protein to an ion channel is stabilized in a state close to the desensitized form of ligand-gated ion channels

To understand the mechanism of allosteric coupling between the ligand-binding domain and the ion channel of the Cys-loop ligand-gated ion channels (LGICs), we fused the soluble acetylcholine-binding protein (AChBP), which lacks an ion channel, to either the cationic serotonin type-3A ion channel (5HT 3A) or the anionic glycine ion channel. Both linear chimeras expressed in HEK-293 cells display high affinity for the nicotinic agonist epibatidine ( K D = 0.2 – 0.5 nM ), but are not targeted to the cell surface. Only after substituting a ring of three loops located at the putative membrane side of the AChBP three-dimensional structure by the homologous residues of 5HT 3A, the resulting chimera AChBP(ring)/5HT 3A ( i) still displayed on intact cells an apparent high affinity for epibatidine, yet with a fourfold decrease ( K D = 2.1 nM ), ( ii) displayed a high proportion of low affinity sites (11 ± 7 μM) for the resting state stabilizing competitive antagonist α-bungarotoxin and ( iii) was successfully targeted to the cell surface, as seen by immunofluorescence labelling. The AChBP(ring)/5HT 3A chimera forms a pentameric structure, as revealed by sucrose gradient sedimentation. However, no whole-cell patch-clamp currents were detectable. Interestingly, binding assays with membrane fragments prepared from cells expressing AChBP(ring)/5HT 3A showed a decrease in the apparent affinity for the agonists nicotine and epibatidine (5-fold), concomitant with an increase in the proportion of high-affinity sites (48 ± 1 nM) for α-bungarotoxin. These results indicate that fusion of AChBP to an ion channel forms a pentameric receptor exposed to the cell surface and able to convert between discrete allosteric states, but stabilized in a high affinity state for epibatidine that likely corresponds to a desensitized form of LGICs. These artificial chimeras might offer a useful system to investigate signal transduction in LGICs. To cite this article: T. Grutter et al., C. R. Biologies 328 (2005).

Opposing Effects of C-Reactive Protein Isoforms on Shear-Induced Neutrophil-Platelet Adhesion and Neutrophil Aggregation in Whole Blood

Plasma C-reactive protein (CRP) level is a powerful predictor of cardiovascular events. However, it is not known whether CRP could affect neutrophil-platelet adhesion and neutrophil aggregation, key events in acute coronary syndromes. Emerging in vitro evidence suggests that some bioactivities of CRP are expressed on loss of the pentameric symmetry, resulting in formation of modified or monomeric CRP (mCRP). We studied the impact of human native CRP and bioengineered mCRP that cannot rearrange into the pentameric structure on the kinetics of neutrophil-platelet adhesion and neutrophil aggregation in whole blood subjected to shear (approximately 100 s(-1)) using real-time flow cytometry. Shear resulted in upregulation of platelet P-selectin expression, leading to platelet capture of neutrophils and subsequent neutrophil aggregation, which was dependent on P-selectin, L-selectin, and CD18. Native CRP at clinically relevant concentrations markedly attenuated these changes. The residual amount of neutrophil adhesion was blocked with anti-CD18 or anti-CD11b antibody. By contrast, mCRP concentration-dependently enhanced shear-induced platelet P-selectin expression and increased the rate and extent of formation of both neutrophil-platelet and neutrophil-neutrophil aggregates. Complete abrogation of platelet-neutrophil adhesion and neutrophil aggregation required both anti-P-selectin and anti-CD18 antibodies but not anti-L-selectin antibody. The CRP action was markedly inhibited by an anti-CD32 antibody, whereas the mCRP effects were significantly attenuated by an anti-CD16 antibody. These results indicate that native CRP inhibits platelet activation and prevents platelet capture of neutrophils, whereas mCRP displays potent prothrombotic activities under low levels of shear. Thus, mCRP rather than native CRP may precipitate acute coronary syndromes.

Discovery of Recurrent Structural Motifs for Approximating Three‐Dimensional Protein Structures

AbstractThe scope of conformation space that protein molecules can adopt is a problem of significant interest. Previous studies by other groups have shown that there are stereochemical constraints that confine local protein structures to a limited range of conformations. Furthermore, the results of many groups have demonstrated that the sequence‐to‐structure relationship remains detectable to some extent on a local level. By studying the conformational space of local protein structures, we may obtain more information concerning the constraints on local structural space and the sequence‐to‐structure mapping, hence facilitate ab initio structure prediction. In this study, we propose a novel algorithm that automatically discovers recurrent pentamer structures in proteins.The algorithm starts by applying Expectation‐Maximization (EM) clustering to the distances between non‐adjacent backbone Cα atoms in a large set of pentamer fragments. A rough partition of the conformation space can thus be derived. In the second stage, by applying a split‐and‐merge algorithm, we can obtain a finite number of clusters and guarantee the homogeneity and distinctiveness of each one. Each cluster of protein structures is represented by a centroid structure. The results show that, with 40 major representative structures, we can approximate most of the protein fragments with an error of 0.378 Å. With only 20 types of structures, the fragment structures can still be modeled at 0.44 Å, which is comparable to or better than the performance of previous methods. We term the representatives “building blocks.” On the global level, we demonstrate that by concatenating different combinations of building blocks, we can model whole protein structures at high resolution: a resolution of 2.54 Å can be achieved simply by using 10 types of building blocks. This finding suggests that the study of molecular structures can be hugely simplified using this reduced representation.

Quantum Mechanical Investigation of Thiourea Adsorption on Ag(111) Considering Electric Field and Solvent Effects

The adsorption of different thiourea species was investigated on the 111 surface of silver considering electric field and solvent effects with the objective of (a) elucidating the nature of the adsorbed species, mainly inan electrochemical environment, and (b) understanding the energetics and mechanisms of the surface reactions, particularly the oxidative adsorption of thiourea. We first considered the adsorption in vacuum of molecular species such as thiourea (TU), canonical thiourea, and formamidine disulfide as well as the adsorption of the canonical thiourea radical resulting from the cleavage of the S-H bond. The molecular adsorption of TU in vacuum has the highest binding energy (-33.2 kcal/mol) when the molecular plane is parallel to the surface. However, the strongest surface bond was observed for the canonical thiourea radical (-42.2 kcal/mol) whose nature is the same as that of the methanethiol radical. Externally applied electric fields perpendicular to the surface have an important effect on the orientation and surface bonding of thiourea, leading to an upright configuration of the molecule and to a strengthening of the sulfur-surface bond. Solvent effects were first considered to investigate the equilibria of thiourea in solution with the purpose of elucidating the nature of the predominant species which, in turn, constitutes the initial adsorbate on the surface. In neutral and basic pHs, the thiourea molecule is the predominant species, whereas in acidic electrolytes, the thiouronium ion readily forms. The microsolvation of thiourea with up to 16 water molecules was investigated to determine the structure of the hydration shell and the interaction energy between TU and water molecules. Water molecules form a compact cage around thiourea characterized by linked tetramer and pentamer structures. The NH 2 groups form hydrogen bonds with water molecules, whereas the sulfur atom is poorly hydrated. The implications of the structure of water around thiourea on the adsorption geometry of the molecule are discussed. The oxidative adsorption mechanism of thiourea resulting in chemisorbed canonical thiourea was investigated considering charge-transfer processes, solvent effects, and the presence of a counterion.

Statistical mechanics model for the interaction between the neurotransmitter gamma-aminobutyric acid and GABAA receptors.

Interactions between the neurotransmitter gamma-aminobutyric acid (GABA) and GABAA receptor ion channels play an important role in the central nervous system. A statistical mechanics model is proposed for the interaction between GABA and GABAA receptors. The model provides good fits to the electrophysiology data as well as an estimation of receptor activation energies, and predicts the temperature dependence consistent with measurements. In addition, the model provides insights into single channel conductance measurements. This model is also applicable to other ligand-gated ion channels with similar pentameric structures.

Conformational rearrangement in C-reactive protein is required for proinflammatory actions on human endothelial cells.

C-reactive protein (CRP) has been suggested to actively amplify the inflammatory response underlying coronary heart diseases by directly activating endothelial cells. In this study, we investigated whether loss of the cyclic pentameric structure of CRP, resulting in formation of modified or monomeric CRP (mCRP), is a prerequisite for endothelial cell activation. We examined the impact of native CRP and mCRP on the production of monocyte chemoattractant protein-1 (MCP-1) and interleukin-8 (IL-8), key regulators of leukocyte recruitment, and on the expression of intercellular adhesion molecule-1 (ICAM-1), E-selectin, and vascular adhesion molecule-1 (VCAM-1) in human cultured coronary artery endothelial cells (HCAECs). Incubation with mCRP for 4 hours increased MCP-1 and IL-8 secretion and mRNA levels and expression of ICAM-1, E-selectin, and VCAM-1 protein and mRNA. Significant induction occurred at 1 to 5 microg/mL, reached a maximum at 30 microg/mL, and did not require the presence of serum. Native CRP was without detectable effects at 4 hours, whereas it enhanced cytokine release after a 24-hour incubation. An anti-FcgammaRIII (CD16) but not an anti-FcgammaRII (CD32) antibody produced a 14% to 32% reduction of the mCRP effects (P<0.05). mCRP but not CRP evoked phosphorylation of p38 mitogen-activated protein kinase, and inhibition of this kinase with SB 203580 reversed the effects of mCRP. Furthermore, culture of HCAECs in the presence of SB203580 markedly decreased mCRP-stimulated E-selectin and ICAM-1-dependent adhesion of neutrophils to HCAECs (P<0.001). Loss of pentameric symmetry in CRP, resulting in formation of mCRP, promotes a proinflammatory HCAEC phenotype through a p38 MAPK-dependent mechanism.

High Order Quaternary Arrangement Confers Increased Structural Stability to Brucella sp. Lumazine Synthase

The penultimate step in the pathway of riboflavin biosynthesis is catalyzed by the enzyme lumazine synthase (LS). One of the most distinctive characteristics of this enzyme is the structural quaternary divergence found in different species. The protein exists as pentameric and icosahedral forms, built from practically the same structural monomeric unit. The pentameric structure is formed by five 18-kDa monomers, each extensively contacting neighboring monomers. The icosahedrical structure consists of 60 LS monomers arranged as 12 pentamers giving rise to a capsid exhibiting icosahedral 532 symmetry. In all lumazine synthases studied, the topologically equivalent active sites are located at the interfaces between adjacent subunits in the pentameric modules. The Brucella sp. lumazine synthase (BLS) sequence clearly diverges from pentameric and icosahedric enzymes. This unusual divergence prompted us to further investigate its quaternary arrangement. In the present work, we demonstrate by means of solution light scattering and x-ray structural analyses that BLS assembles as a very stable dimer of pentamers, representing a third category of quaternary assembly for lumazine synthases. We also describe by spectroscopic studies the thermodynamic stability of this oligomeric protein and postulate a mechanism for dissociation/unfolding of this macromolecular assembly. The higher molecular order of BLS increases its stability 20 degrees C compared with pentameric lumazine synthases. The decameric arrangement described in this work highlights the importance of quaternary interactions in the stabilization of proteins.

Modification of the cholera toxin B subunit coding sequence to enhance expression in plants

The cholera toxin B subunit (CTB) contains five identical polypeptides and targets glycosphingolipid receptors on eukaryotic cell surfaces. Increased expression of CTB in plants is critical for the development of edible vaccines. In this study, the coding sequence of the CTB gene was optimized, based on the modification of codon usage to that of tobacco plant genes and the removal of mRNA-destabilizing sequences. The synthetic CTB gene was cloned into a plant expression vector and expressed in tobacco plants under the control of the CaMV 35S promoter. The recombinant CTB protein constituted approximately 1.5% of the total soluble protein in transgenic tobacco leaves. This level of CTB production was approximately 15-fold higher than that in tobacco plants that were transformed with the bacterial CTB gene. The recombinant CTB produced by tobacco plants demonstrated strong affinity for GM1-ganglioside, which indicates that the sites required for binding and proper folding of the pentameric CTB structure were conserved. This is the first report on the optimization of the CTB-coding sequence to give a dramatic increase in CTB expression in plants.

Thermodynamics of forming water clusters at various temperatures and pressures by Gaussian-2, Gaussian-3, complete basis set-QB3, and complete basis set-APNO model chemistries; implications for atmospheric chemistry.

The Gaussian-2, Gaussian-3, complete basis set- (CBS-) QB3, and CBS-APNO methods have been used to calculate Delta H degrees and Delta G degrees values for neutral clusters of water, (H(2)O)(n), where n = 2-6. The structures are similar to those determined from experiment and from previous high-level calculations. The thermodynamic calculations by the G2, G3, and CBS-APNO methods compare well against the estimated MP2(CBS) limit. The cyclic pentamer and hexamer structures release the most heat per hydrogen bond formed of any of the clusters. While the cage and prism forms of the hexamer are the lowest energy structures at very low temperatures, as temperature is increased the cyclic structure is favored. The free energies of cluster formation at different temperatures reveal interesting insights, the most striking being that the cyclic trimer, cyclic tetramer, and cyclic pentamer, like the dimer, should be detectable in the lower troposphere. We predict water dimer concentrations of 9 x 10(14) molecules/cm(3), water trimer concentrations of 2.6 x 10(12) molecules/cm(3), tetramer concentrations of approximately 5.8 x 10(11) molecules/cm(3), and pentamer concentrations of approximately 3.5 x 10(10) molecules/cm(3) in saturated air at 298 K. These results have important implications for understanding the gas-phase chemistry of the lower troposphere.

Modeling the structure of the respiratory syncytial virus small hydrophobic protein by silent‐mutation analysis of global searching molecular dynamics

Human respiratory syncytial virus (RSV) encodes a small hydrophobic (SH) protein, whose function in the life cycle of the virus is unknown. Recent channel activity measurements of the protein suggest that like other viroporins, SH may assemble into a homo-oligomeric ion channel. To further our understanding of this potentially important protein, a new strategy was implemented in order to model the transmembrane oligomeric bundle of the protein. Global searching molecular dynamic simulations of SH proteins from eight different viral strains, each at different oligomeric states, as well as different lengths of the putative transmembrane domain, were undertaken. Taken together, a total of 45 different global molecular dynamic simulations pointed to a single pentameric structure for the protein that was found in all of the different variants. The model of the structure obtained is a channel-like homopentamer whose minimal transmembrane pore diameter is 1.46 A.