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1,3-dicholoro-1,3-bis(dialkylamino)propenium salts are prepared by condensing N,N,-dialkylamides with dichloromethylene dimethyliminium chloride ("Viehe's salt"). The products are versatile synthons and can be used to make a variety of lig-and types. Here, we report the syntheses of a number of new propenium salts and provide comprehensive characterization data for some that have already been described. Additionally we have prepared a series of related 1,3-dihalo-1,3-bis(dimethylamino)propenium salts via halogen exchange reactions (F through I). Finally all of the compounds described, including Viehe's Salt and several hydrolysis products, have been characterized in the solid state using single crystal X-ray diffraction. The symmetrical cations [[[Me.sub.2]NC(X)CHC(X)N[Me.sub.2]].sup.+] were found to take one of two general structural forms. The first (X = H or F) is planar, presumably maximizing the delocalization within the backbone of the cation, while the second form is markedly twisted (X = Cl, Br or I). Calculations indicate that the latter develop increasingly positive regions of the electrostatic potential on the halide substituents, allowing cation/anion interactions via halogen bonding to dominate in the extended solids. The results of these studies illustrate the importance of the differing interionic interactions in a homologous series of small organic salts. Key words: X-ray crystallography, carbocation, 1,3-bis(dimethylamino)propenium cation, vinamidinium ion, Viehe's salt, Introduction 1,3-Bis(dimethylamino)propenium ions, or vinamidinium ions, have been shown to be powerful synthons for the preparation of [C.sub.3]-backbone ligands, since they are easily functionalized and readily undergo substitution. In the [...]

(Bromoethynyl)benzene dissolved in hexane reacts with l,3-bis(2,4,6-trimethylphenyl)imidazol-2-ylidene (IMes) to produce the acetylenic amidinium salt [Ph-C[equivalent to]C-IMes][Br]. Treatment of the bromide salt with [KPF.sub.6] then results in the isolation of solid [Ph-C[equivalent to]C-IMes][[PF.sub.6]]. Both salts have been characterized spectroscopically and their structures determined using X-ray crystallography. Crystals of a hydrolysis product were also isolated from this reaction on one occasion and its structure is reported. This led us to explore the reactivity of the bromide salt further. It was found to react rapidly with sodium azide (Na[N.sub.3]), azidotrimethylsilane ([(C[H.sub.3]).sub.3]Si-[N.sub.3]), cyanotrimethylsilane ([(C[H.sub.3]).sub.3]Si-CN), and also with IMes. All of the addition products have been characterized using X-ray crystallography. Key words: X-ray crystallography, alkyne, N-heterocyclic carbene, C-H bond activation, triazole, Introduction Acetylenic iminium 1 and amidinium 2 (Fig. 1) ions are interesting species since they combine, in one molecule, two reactive and versatile functional groups in close proximity. It was [...]

The synthesis and characterisation (IR, NMR, HR-MS, and X-ray crystallography) of six oxazoline Zn complexes, containing enolate ligands derived from a series of (Z)-1-R-2-(4',4'-dimethyl-2'-oxazolin-2'-yl)-efh-1-en-1-ols (R=[C.sub.6][H.sub.5], [C.sub.6][H.sub.4]-p-N[O.sub.2], [C.sub.6][H.sub.4]-p-O[CH.sub.3], [CF.sub.3], [CH.sub.3], or [CMe.sub.3]) is detailed. All derivatives are air-stable solids of formula Zn[([L.sup.n]).sub.2] and are formed in the presence of the title ligands and [NEt.sub.3]. Reasonable yields (53%-82%) are obtained with the exception of R= [CMe.sub.3] (9%); in this case, by-product formation and high solubility affected complex isolation. Crystallographic characterisation reveals that in all cases the ligands are bound in a [kappa].sup.2]-N, O-bonding motif. Little structural diversity is noted within the series; these complexes are further placed into structural context with related Zn materials containing oxazoline, phenolate, and (or) Schiff base ligands. Key words: oxazoline, coordination complexes, zinc, enolate, X-ray crystal structure, Introduction The naturally occurring Group 12 elements hold a unique position within coordination chemistry due to the overwhelming predominance of the +2 oxidation state. This is, of course, a result [...]

The photophysical properties of four 1,3,6-trisubstituted pyrazolo[3,4-d][1,2,3]triazoles were studied by a combination of UV--vis and fluorescence spectroscopy as well as density functional theory (DFT) and time-dependent DFT calculations. All four compounds are weakly fluorescent. Upon protonation with trifluoroacetic acid, all compounds display a drastic luminescence turn-on. The addition of excess trifluoroacetic acid further enhances the luminescence intensity, which is tentatively attributed to the formation of extended hydrogen bonding network rather than the formation of doubly protonated 1,3,6-trisubstituted pyrazolo[3,4-d][1,2,3]triazoles. All new compounds were characterized by nuclear magnetic resonance and X-ray crystallography. Key words: luminescence, UV--vis spectroscopy, pyrazole, triazole, fused N-heterocycles, Introduction Nitrogen-rich fused heterocycles are an important class of compounds with wide applications in the rapidly expanding fields of medicinal chemistry, drug discovery, and functional material development. (1-8) Pyrazolo[3,4-d][1,2,3]triazoles are [...]

Five Cu(II) hydroxo-bridged complexes of the type [[Cu.sub.2][(μ-OH).sub.2][(L).sub.2]][[A].sub.x] x γ[H.sub.2]O (where L = 1,10-phenanthroline, A = [NCS.sup.-], x = 2, y = 2 (1); L = 2,2'-bipyridine, A = [NCS.sup.-], x = 2, γ = 1 (2); L = 2,2'-bipyridine, A = [Au[(CN).sub.4]]-, x = 2, y = 0 (3) and 2 (4); L = N,N,N',N'-teteramethylethylenediamine, A = [[Pt[(SCN).sub.4]].sup.2-], x = 1, y = 0 (5)) have been prepared and have been characterized by elemental analysis, IR and Raman spectroscopy, X-ray crystallography, and SQUID magnetometry. Complexes 1 and 2 consist of five-coordinate Cu(II) hydroxo-bridged dinuclear units bound to two NCS- anions in the apical positions, 3 and 4 are pseudopolymorphs of a Cu(II) hydroxo-bridged dimer core with [Au[(CN).sub.4]]- counteranions, and 5 is a coordination polymer of Cu(II) hydroxo-bridged dimers with trans-bridging [[Pt[(SCN).sub.4]].sup.2-] anions coordinated in the apical position of the five-coordinate Cu(II) centres. The magnetic susceptibility versus temperature for each system was measured, fit to obtain J-coupling values (which range from -0.37 to 17.8 [cm.sup.-1]), and qualitatively compared to known magnetostructural correlation parameters. Key words: copper, hydroxide, coordination polymers, solid-state chemistry, isothiocyanate, X-ray structure, gold, cyanide. Cinq complexe du Cu(II) avec ponts hydroxydes, du type [[Cu.sub.2][(μ-OH).sub.2][(L).sub.2]][[A].sub.x] x γ[H.sub.2]O (ou L=1,10-phenanthroline, A = [NCS.sup.-], x = 2, y =2 (1); L = 2,2'-bipyridine, A = [NCS.sup.-], x = 2, y = 1 (2); L = 2,2'-bipyridine, A = [[Au[(CN).sub.4]].sup.-] x = 2, y = 0 (3) et 2 (4); L = N,N,N',N'-tetramethylethylenediamine, A = [[Pt[(SCN).sub.4]].sup.2-], x = 1, y =0 (5)), ont ete prepares et caracterises par analyse elementaire, spectroscopie IR et Raman, cristallographie aux rayons X et magnetometrie SQUID. Les complexes 1 et 2 sont formes d'unites binucleaires pentacoordonnees de Cu(II) avec ponts hydroxydes, reliees a deux anions NCS- situes en positions apicales, 3 et 4 sont pseudopolymorphes d'un noyau dimerique de Cu(II) a ponts hydroxydes comportant des contranions [[Au[(CN).sub.4]].sup.-] et 5 est un polymere de coordination de dimeres de Cu(II) a ponts hydroxydes formes d'anions pontants [[Pt[(SCN).sub.4]].sup.2-] en position trans, coordonnes en position apicale des centres Cu(II) pentacoordonnes. La susceptibilite magnetique et la temperature ont ete mesurees pour chaque systeme, de maniere a obtenir les valeurs de couplage/ (comprises entre -0,37 et 17,8 [cm.sup.- 1]), et quantitativement comparees aux parametres connus de correlation magnetostructurale. [Traduit par la Redaction] Mot-cles: cuivre, hydroxyde, polymeres de coordination, chimie de l'etat solide, isothiocyanate, structure aux rayons X, or, cyanure., Introduction The structural and magnetic aspects of dinuclear Cu(II) hydroxobridged complexes have been of strong interest to the magneto-chemistry community because their [d.sup.9]-[d.sup.9] configuration allows for a wide range of [...]

The tetranuclear neutral species [[[Cr.sup.III.sub.4][([μ.sub.3]-O).sub.2][([μ.sub.2]- C[H.sub.3]C[O.sub.2]).sub.7]([sup.t][bpy.sup.0])([sup.t][bpy.sup.*])].sup.0] x 3THF has been prepared from a mixture of [[Cr.sup.II.sub.2][([μ.sub.2]-C[H.sub.3]C[O.sub.2]).sub.4]] x 2[H.sub.2]O and 4,4'-di-tert- butyl-2,2'-bipyridine (1:2) in tetrahydrofuran (THF) under anaerobic conditions. Black crystals of [[[Cr.sup.III.sub.4][(μ-O).sub.2][(μ- C[H.sub.3]C[O.sub.2]).sub.7][([sup.t]bpy).sub.2].sup.0] were grown and investigated by X-ray crystallography: a [[[Cr.sub.4][O.sub.2]].sup.8+] butterfly core structure has been identified. In addition, the presence of seven [μ.sub.2]-acetate bridges, a neutral ([sup.t][bpy.sup.0]) ligand, and a single π-radical anion [([sup.t][bpy.sup.*]).sup.1-] have been clearly identified. The magnetic properties have been investigated by solid-state SQUID (superconducting quantum interference device) measurements (2-300 K) and a coupling model is presented. Its X-band electron paramagnetic resonance spectrum has been recorded at 10 and 300 K in toluene solution and successfully simulated. The spin of the [([sup.t][bpy.sup.*]).sup.1-]radical couples strongly antiferromagnetically to one Cr(III) center ([J.sub.4] = -129 [cm.sup.-1]). These properties are compared with those reported previously for the monocationic species [[[Cr.sup.III.sub.4][([μ.sub.3]-O).sub.2] [([μ.sub.2]-C[H.sub.3]C[O.sub.2]).sub.7][([bpy.sup.0]).sub.2](P[F.sub.6]) in Bino et al. (Inorg. Chem. 1991, 30, 856). Key words: tetranuclear chromium(III) complex, electronic structure, electron spin resonance spectroscopy, ligand π-radical, molecular magnetochemistry. Resume : L'espece tetranucleaire neutre, de formule [[[Cr.sup.III.sub.4][([[my].sub.2]- O).sub.2][([[my].sub.2]- C[H.sub.3]C[O.sub.2]).sub.7]([sup.t][bpy.sup.0])([sup.t][bpy.sup.*])].sup.0] x 3THF, a ete preparee a partir d'un melange de [[Cr.sup.II.sub.2][([μ.sub.2]-C[H.sub.3]C[O.sub.2]).sub.4]] x 2[H.sub.2]O et de 4,4'- di-tert-butyl-2,2'- bipyridine (1:2) dans du tetrahydrofuranne (THF) et en conditions anaerobie. Des cristaux noirs de [[[Cr.sup.III.sub.4][(μ-O).sub.2][(μ- C[H.sub.3]C[O.sub.2]).sub.7][([sup.t]bpy).sub.2].sup.0] se sont formes et ont ete analyses par cristallographie aux rayons X. Une structure centrale de formule [[[Cr.sub.4][O.sub.2]].sub.8+] a ete alors identifiee. De plus, la presence de sept ponts [[my].sub.2]-acetate, un ligand neutre, ([sup.t][bpy.sup.0]), et d'un anion π-radical seul [([sup.t][bpy.sup.*]).sup.1-] a ete clairement detectee. Les proprietes magnetiques du complexe neutre ont ete etudiees en effectuant des mesures a l'etat solide avec un SQUID (<< superconducting quantum interference device >>) (entre 2 et 300 K) et un modele de couplage propose. Son spectre resonance paramagnetique electronique en bande X a ete mesure a 10 et 300 K dans une solution de toluene et simule avec succes. On observe un fort couplage antiferromagnetique entre le spin du radical [([sup.t][bpy.sup.*]).sup.1-] et un centre Cr(III)([J.sub.4] = -129 [cm.sup.-1]). Ces proprietes sont compares a celles observees precedemment dans le cas des especes monocationiques [[[Cr.sup.III.sub.4][([μ.sub.3]-O).sub.2] [([μ.sub.2]-C[H.sub.3]C[O.sub.2]).sub.7][([bpy.sup.0]).sub.2](P[F.sub.6]) (Bino et al. Inorg. Chem. 1991, 30, 856). [Traduit par la Redaction] Mots-cles : complexe chrome(III) tetranucleaire, structure electronique, spectroscopie par resonnance de spin electronique, ligand π-radical, magnetochimie moleculaire., Introduction The synthesis, the molecular structure and the spectroscopic (UV-vis, near-infrared, etc.) properties as well as the most interesting molecular magnetism of a series of isostructural tetranuclear complexes of the [...]

The use of three shape-persistent, conjugated macrocycles 2a-2c as ligands for self-assembly reactions is described. The macrocycles have been characterized through a combination of spectroscopic analyses and, for compounds 2b and 2c, X-ray crystallographic analysis. Whereas the reaction of 2a with the cis-Pt(n) species 3 successfully provides the porous solid 4a, the analogous reaction of 2b and 2c with 3 leads only to mixtures of products. The application of continuous- flow hyperpolarized [sup.129]Xe NMR spectroscopy to investigate the solid-state pores of macrocycles 2b and 2c and the supramolecular complex 4a as a function of temperature is described. All three species show permanent porosity upon removal of co-crystallized solvent molecules. Using trends in the [sup.129]Xe chemical shifts and temperature-dependent dynamics of Xe atoms in the solids, information is obtained on the nature of the pores in these systems. Using the [sup.129]Xe NMR data for complex 4a, the effective heat of adsorption (Δ[H.sub.ads]) was calculated to be ~29 kJ [mol.sup.-1]. Key words: macrocycles, solid-state NMR, supramolecular chemistry, xenon NMR, porous solids, metal-organic framework. On decrit l'utilisation des trois macrocycles conjugues a forme persistante, 2a-2c, comme ligands pour des reactions d'auto-assemblage. On a caracterise les macrocycles par une combinaison d'analyses spectroscopiques et, dans le cas des composes 2b et 2c, par analyse de donnees cristallographiques des rayons-X. Alors que la r?action du compose 2a avec l'espece cis-Pt(n), 3, conduit avec succes a la formation du solide poreux 4a, les reactions analogues des composes 2b et 2c avec le compose 3 ne conduisent qu'a des melanges de produits. On decrit l'application de la spectroscopie RMN du [sup.129]Xe en flux continu hyperpolarise a l'etude des pores a l'etat solide des macrocycles 2b et 2c et du complexe supramol?culaire 4a en fonction de la temperature. Les trois especes presentent chacune une porosite permanente lors de l'elimination des molecules de solvant cocristallisees. En se basant sur les tendances dans les deplacements chimiques du [sup.129]Xe et sur la variation de la dynamique des atomes de xenon en fonction de la temperature, on a obtenu des informations sur la nature des pores dans ces syst?mes. En utilisant les donnees de la RMN du [sup.129]Xe pour le complexe 4a, on a determine que la chaleur effective d'adsorption (Δ[H.sub.ads]) est approximativement egale a 29 kJ [mol.sup.-1]. Mots-cles : macrocycles, RMN a l'etat solide, chimie supramol?culaire, RMN du [sup.129]Xe, solides poreux, squelette organometallique. [Traduit par la R?daction], Introduction The synthesis of organic and organometallic porous materials is a rapidly expanding field of synthetic chemistry. (1-5) Recent efforts to use organic and metal-organic macrocycles to form ordered materials [...]

Chikungunya virus (CHIKV) is an emerging mosquito-borne alphavirus that has caused widespread outbreaks of debilitating human disease in the past five years (1). CHIKV invasion of susceptible cells is mediated by two viral glycoproteins, E1 and E2, which carry the main antigenic determinants and form an icosahedral shell at the virion surface. Glycoprotein E2, derived from furin cleavage of the p62 precursor into E3 and E2, is responsible for receptor binding, and E1 for membrane fusion. In the context of a concerted multidisciplinary effort to understand the biology of CHIKV (2), here we report the crystal structures of the precursor p62-E1 heterodimer and of the mature E3-E2-E1 glycoprotein complexes. The resulting atomic models allow the synthesis of a wealth of genetic, biochemical, immunological and electron microscopy data accumulated over the years on alphaviruses in general. This combination yields a detailed picture of the functional architecture of the 25 MDa alphavirus surface glycoprotein shell. Together with the accompanying report on the structure of the Sindbis virus E2-E1 heterodimer at acidic pH (ref. 3), this work also provides new insight into the acid-triggered conformational change on the virus particle and its inbuilt inhibition mechanism in the immature complex., Although the symptoms of the disease were first described in the eighteenth century (4), CHIKV was first isolated only in 1952 during a dengue outbreak in Tanzania (5). The name [...]

Firefly luciferase (FLuc), an ATP-dependent bioluminescent reporter enzyme, is broadly used in chemical biology and drug discovery assays. PTC124 (Ataluren; (3-[5-(2-fluorophenyl)-1,2,4-oxadiazol-3-yl] benzoic acid) discovered in an FLuc-based assay targeting nonsense codon suppression, is an unusually potent FLuc-inhibitor. Paradoxically, PTC124 and related analogs increase cellular FLuc activity levels by posttranslational stabilization. In this study, we show that FLuc inhibition and stabilization is the result of an inhibitory product formed during the FLuc-catalyzed reaction between its natural substrate, ATP, and PTC124. A 2.0 [Angstrom] cocrystal structure revealed the inhibitor to be the acyI-AMP mixed-anhydride adduct PTC124-AMP, which was subsequently synthesized and shown to be a high-affinity multisubstrate adduct inhibitor (MAI; [K.sub.D] = 120 pM) of FLuc. Biochemical assays, liquid chromatography/mass spectrometry, and near-attack conformer modeling demonstrate that formation of this novel MAI is absolutely dependent upon the precise positioning and reactivity of a key meta-carboxylate of PTC124 within the FLuc active site. We also demonstrate that the inhibitory activity of PTC124-AMP is relieved by free coenzyme A, a component present at high concentrations in luciferase detection reagents used for cell-based assays. This explains why PTC124 can appear to increase, instead of inhibit, FLuc activity in cell-based reporter gene assays. To our knowledge, this is an unusual example in which the 'off-target' effect of a small molecule is mediated by an MAI mechanism. Ataluren | multisubstrate adduct inhibitor | protein stability | reporter gene assay | X-ray crystallography doi: 10.1073/pnas.0909141107

Photosynthesis makes use of sunlight to convert carbon dioxide into useful biomass and is vital for life on Earth. Crucial components for the photosynthetic process are antenna proteins, which absorb light and transmit the resultant excitation energy between molecules to a reaction centre. The efficiency of these electronic energy transfers has inspired much work on antenna proteins isolated from photosynthetic organisms to uncover the basic mechanisms at play (1-5). Intriguingly, recent work has documented (6-8) that light-absorbing molecules in some photosynthetic proteins capture and transfer energy according to quantum-mechanical probability laws instead of classical laws (9) at temperatures up to 180 K. This contrasts with the long-held view that long-range quantum coherence between molecules cannot be sustained in complex biological systems, even at low temperatures. Here we present two-dimensional photon echo spectroscopy (10-13) measurements on two evolutionarily related light-harvesting proteins isolated from marine cryptophyte algae, which reveal exceptionally long-lasting excitation oscillations with distinct correlations and anti-correlations even at ambient temperature. These observations provide compelling evidence for quantumcoherent sharing of electronic excitation across the 5-nm-wide proteins under biologically relevant conditions, suggesting that distant molecules within the photosynthetic proteins are 'wired' together by quantum coherence for more efficient light-harvesting in cryptophyte marine algae., Cryptophytes are eukaryotic algae that live in marine and freshwater environments. They are members of an evolutionary group notable because their photosynthetic apparatus was acquired from red algae by a [...]

In an effort to find new pharmacological modalities to overcome resistance to ATP-binding-site inhibitors of Bcr-Abl, we recently reported the discovery of GNF-2, a selective allosteric Bcr-Abl inhibitor. Here, using solution NMR, X-ray crystallography, mutagenesis and hydrogen exchange mass spectrometry, we show that GNF-2 binds to the myristate-binding site of Abl, leading to changes in the structural dynamics of the ATP-binding site. GNF-5, an analogue of GNF-2 with improved pharmacokinetic properties, when used in combination with the ATP-competitive inhibitors imatinib or nilotinib, suppressed the emergence of resistance mutations in vitro, displayed additive inhibitory activity in biochemical and cellular assays against T315I mutant human Bcr-Abl and displayed in vivo efficacy against this recalcitrant mutant in a murine bone-marrow transplantation model. These results show that therapeutically relevant inhibition of Bcr-Abl activity can be achieved with inhibitors that bind to the myristate-binding site and that combining allosteric and ATP- competitive inhibitors can overcome resistance to either agent alone., Chronic myelogenous leukaemia (CML) is a haematological malignancy caused by a chromosomal rearrangement that generates a fusion protein, Bcr-Abl, with deregulated tyrosine kinase activity. Although clinical remission is usually achieved [...]

Preorganization is shown to endow a protein with extraordinary conformational stability. This preorganization is achieved by installing side-chain substituents that impose stereoelectronic and steric effects that restrict main-chain torsion angles. Replacing proline residues in [(ProProGly).sub.7] collagen strands with 4-fluoroproline and 4-methylproline leads to the most stable known triple helices, having [T.sub.m] values that are increased by >50[degrees]C Differential scanning calorimetry data indicate an entropic basis to the hyperstability, as expected from an origin in preorganization. Structural data at a resolution of 1.21 [Angstrom] reveal a prototypical triple helix with insignificant deviations to its main chain, even though 2/3 of the residues are nonnatural. Thus, preorganization of a main chain by subtle changes to side chains can confer extraordinary conformational stability upon a protein without perturbing its structure. collagen triple helix | nonnatural amino acid | preorganization | protein stability | x-ray crystallography www.pnas.org/cgi/doi/10.1073/pnas.0909592107

Problem statement: Finding an accurate RNA structural alignment from primary sequence due to it is time consuming and computationally NP-hard problem is a major bioinformatics challenge. According to our investigation majority of current researches were concerned on achieving faster execution time, improving space complexity and better cache management. Recently one research introduced cache-efficient Chip Multiprocessor (CMP) algorithms with good speed-up to exploit parallelism in detection the critical path length. Our contribution in this article was a comprehensive survey of methods for solving RNA secondary structure prediction with Pseudoknots (PK) and sequence alignment in bioinformatics. The aim was to highlight the challenges related issues which would provide sufficient information to assist the new coming researchers in this field as well as a good reference guide for bioinformatics professionals. Approach: We computed various algorithms that predicted an RNA molecules secondary structure from primary sequence, without pseudoknots from one side and pseudoknotted RNA secondary structure in the other side. Furthermore, we also reviewed and compared in two tables the methods that developed for RNA structural predictions. Results: Our findings of this survey confirmed that Dynamic Programming (DP) method via CMP algorithms can be used to predict the RNA secondary structure with simple PK and it gives good results. Conclusion: The methods for predicting RNA's structural are coming in two groups: Firstly, pseudoknotted RNA structural problem is computationally complex and secondly, common methods significantly gave not accurate enough results for predicting pseudoknotted RNA. Key words: Bioinformatics, RNA secondary structure, pseudoknots, dynamic programming, NP-complete, INTRODUCTION Bioinformatics is a computer application to manage the biological information and it uses computer to gather, store, analyze, manipulate, interpret and integrate biological, genetic information and macromolecules (Deoxyribo Nucleic [...]

The proper glycosylation of proteins trafficking through the Golgi apparatus depends upon the conserved oligomeric Golgi (COG) complex. Defects in COG can cause fatal congenital disorders of glycosylation (CDGs) in humans. The recent discovery of a form of CDG, caused in part by a COG4 missense mutation changing Arg 729 to Trp, prompted us to determine the 1.9 [Angstrom] crystal structure of a Cog4 C-terminal fragment. Arg 729 is found to occupy a key position at the center of a salt bridge network, thereby stabilizing Cog4's small C-terminal domain. Studies in HeLa cells reveal that this C-terminal domain, while not needed for the incorporation of Cog4 into COG complexes, is essential for the proper glycosylation of cell surface proteins. We also find that Cog4 bears a strong structural resemblance to exocyst and Dsl1p complex subunits. These complexes and others have been proposed to function by mediating the initial tethering between transport vesicles and their membrane targets; the emerging structural similarities provide strong evidence of a common evolutionary origin and may reflect shared mechanisms of action. congenital disorder of glycosylation | Golgi apparatus multi-subunit tethering complex | vesicle trafficking | X-ray crystallography

A detailed single-crystal X-ray study of conformationally flexible sulfonimide-based dendritic molecules with systematically varied molecular architectures was undertaken. Thirteen crystal structures reported in this work include 9 structures of the second-generation dendritic sulfonimides decorated with different aryl groups, 2 compounds bearing branches of both second and first generation, and 2 representatives of the first generation. Analysis of the packing patterns of 9 compounds bearing second-generation branches shows that despite their lack of strong directive functional groups there is a repeatedly reproduced intermolecular interaction mode consisting in an anchor-type packing of complementary second-generation branches of neighbouring molecules. The observed interaction tolerates a wide range of substituents in meta- and para-positions of the peripheral arylsulfonyl rings. Quantum chemical calculations of the molecule-molecule interaction energies agree at the qualitative level with the packing preferences found in the crystalline state. The calculations can therefore be used as a tool to rationalize and predict molecular structures with commensurate and non-commensurate branches for programming of different packing modes in crystal. dendrimers | single-crystal X-ray | sulfonimides | supramolecular chemistry

Tryptophan biosynthesis is subject to exquisite control in species of Bacillus and has become one of the best-studied model systems in gene regulation. The protein TRAP (trp RNA-binding attenuation protein) predominantly forms a ring-shaped 11-mer, which binds cognate RNA in the presence of tryptophan to suppress expression of the trp operon. TRAP is itself regulated by the protein Anti-TRAP, which binds to TRAP and prevents RNA binding. To date, the nature of this interaction has proved elusive. Here, we describe mass spectrometry and analytical centrifugation studies of the complex, and 2 crystal structures of the TRAP--Anti-TRAP complex. These crystal structures, both refined to 3.2-[Angstrom] resolution, show that Anti-TRAP binds to TRAP as a trimer, sterically blocking RNA binding. Mass spectrometry shows that 11-mer TRAP may bind up to 5 AT trimers, and an artificial 12-mer TRAP may bind 6. Both forms of TRAP make the same interactions with Anti-TRAP. Crystallization of wild-type TRAP with Anti-TRAP selectively pulls the 12-mer TRAP form out of solution, so the crystal structure of wild-type TRAP--Anti-TRAP complex reflects a minor species from a mixed population. protein complex | X-ray crystallography | transcription regulation | analytical ultracentrifugation | mass spectrometry

Although x-ray crystallography is the most widely used method for macromolecular structure determination, it does not provide dynamical information, and either experimental tricks or complementary experiments must be used to overcome the inherently static nature of crystallographic structures. Here we used specific x-ray damage during temperature-controlled crystallographic experiments at a third-generation synchrotron source to trigger and monitor (Shoot-and-Trap) structural changes putatively involved in an enzymatic reaction. In particular, a nonhydrolyzable substrate analogue of acetylcholinesterase, the 'off-switch' at cholinergic synapses, was radiocleaved within the buried enzymatic active site. Subsequent product clearance, observed at 150 K but not at 100 K, indicated exit from the active site possibly via a 'backdoor.' The simple strategy described here is, in principle, applicable to any enzyme whose structure in complex with a substrate analogue is available and, therefore, could serve as a standard procedure in kinetic crystallography studies. acetylcholinesterase | kinetic crystallography | structure-dynamics-function relationships | energy landscape | synchrotron radiation

The hemagglutinin-esterases (HEs) are a family of viral envelope glycoproteins that mediate reversible attachment to O-acetylated sialic acids by acting both as lectins and as receptor-destroying enzymes (RDEs). Related HEs occur in influenza C, toro-, and coronaviruses, apparently as a result of relatively recent lateral gene transfer events. Here, we report the crystal structure of a coronavirus (CoV) HE in complex with its receptor. We show that CoV HE arose from an influenza C-like HE fusion protein (HEF). In the process, HE was transformed from a trimer into a dimer, whereas remnants of the fusion domain were adapted to establish novel monomer-monomer contacts. Whereas the structural design of the RDE-acetylesterase domain remained unaltered, the HE receptor-binding domain underwent remodeling to such extent that the ligand is now bound in opposite orientation. This is surprising, because the architecture of the HEF site was preserved in influenza A HA over a much larger evolutionary distance, a switch in receptor specificity and extensive antigenic variation notwithstanding. Apparently, HA and HEF are under more stringent selective constraints than HE, limiting their exploration of alternative binding-site topologies. We attribute the plasticity of the CoV HE receptor-binding site to evolutionary flexibility conferred by functional redundancy between HE and its companion spike protein S. Our findings offer unique insights into the structural and functional consequences of independent protein evolution after interviral gene exchange and open potential avenues to broad-spectrum antiviral drug design. sialic acid | x-ray crystallography | glycoprotein | lectin | nidovirus

The vacuole-type ATPases (V-ATPases) are proton pumps in various intracellular compartments of eukaryotic cells. Prokaryotic V-ATPase of Enterococcus hirae, closely related to the eukaryotic enzymes, provides a unique opportunity to study ion translocation by V-ATPases because it transports [Na.sup.+] ions, which are easier to detect by x-ray crystallography and radioisotope experiments. The purified rotor ring (K-ring) of the E. hirae V-ATPase binds one [Na.sup.+] ion per K-monomer with high affinity, which is competitively inhibited by [Li.sup.] or [H.sup.+], suggesting that the K-ring can also bind these ions. This finding is also supported by the K-ring structure at 2.8 [Angstrom] in the presence of [Li.sup.+]. Association and dissociation rates of the [Na.sup.+] to and from the purified K-ring were extremely slow compared with the [Na.sup.+] translocation rate estimated from the enzymatic activity, strongly suggesting that interaction with the stator subunit (I-subunit) is essential for [Na.sup.+] binding to/release from the K-ring. membrane protein | rotary motor | x-ray crystallography | lithium ion Enterococcus hirae

Recent advances in cell-free expression protocols have opened a new avenue toward high-resolution structural investigations of membrane proteins by x-ray crystallography and NMR spectroscopy. One of the biggest challenges for liquid-state NMR-based structural investigations of membrane proteins is the significant peak overlap in the spectra caused by large line widths and limited chemical shift dispersion of [alpha]-helical proteins. Contributing to the limited chemical shift dispersion is the fact that [approximately equal to] 60% of the amino acids in transmembrane regions consist of only six different amino acid types. This principle disadvantage, however, can be exploited to aid in the assignment of the backbone resonances of membrane proteins; by [sup.15]N/[sup.13]C-double-labeling of these six amino acid types, sequential connectivities can be obtained for large stretches of the transmembrane segments where number and length of stretches consisting exclusively of these six amino acid types are enhanced compared with the remainder of the protein. We show by experiment as well as by statistical analysis that this labeling scheme provides a large number of sequential connectivities in transmembrane regions and thus constitutes a tool for the efficient assignment of membrane protein backbone resonances. cell-free expression | NMR | stable isotope labeling | protein structure analysis | peak overlap

The externally coordinated arylallyllithiums with internally coordinated arylallyllithiums are compared by using a combination of X-ray and NMR studies. The decrease in barriers is correlated with a decrease in separation of lithium from an allyl terminal carbon.

A new method is described for joint interleaved recording, real-time processing, and analysis of NMR data sets. This new method has removed the most critical time bottlenecks in data acquisition and has increased the value of hyperdimensional NMR spectroscopy in structural biology.

He I photoelectron (PE) spectroscopy is employed to characterize [3] and [5] ladderanes obtained by way of template-controlled syntheses conducted in the organic solid state. The results provide a first correlation with X-ray crystallographic structure data.

The biosynthesis of rebeccamycin, an antitumor compound, involves the remarkable eight-electron oxidation of chlorinated chromopyrrolic acid. Although one rebeccamycin biosynthetic enzyme is capable of generating low levels of the eight-electron oxidation product on its own, a second protein, RebC, is required to accelerate product formation and eliminate side reactions. However, the mode of action of RebC was largely unknown. Using crystallography, we have determined a likely function for RebC as a flavin hydroxylase, captured two snapshots of its dynamic catalytic cycle, and trapped a reactive molecule, a putative substrate, in its binding pocket. These studies strongly suggest that the role of RebC is to sequester a reactive intermediate produced by its partner protein and to react with it enzymatically, preventing its conversion to a suite of degradation products that includes, at low levels, the desired product. flavin enzymes | x-ray crystallography | indolocarbazoles | antitumor

TBP-associated factor 4 (TAF4), an essential subunit of the TFIID complex acts as a coactivator for multiple transcriptional regulators, including Sp1 and CREB. However, little is known regarding the structural properties of the TAF4 subunit that lead to the coactivator function. Here, we report the crystal structure at 2.0-[Angstrom] resolution of the human TAF4-TAFH domain, a conserved domain among all metazoan TAF4, TAF4b, and ETO family members. The hTAF4-TAFH structure adopts a completely helical fold with a large hydrophobic groove that forms a binding surface for TAF4 interacting factors. Using peptide phage display, we have characterized the binding preference of the hTAF4-TAFH domain for a hydrophobic motif, D[PSI][PSI][zeta][zeta][PSI][PHI], that is present in a number of nuclear factors, including several important transcriptional regulators with roles in activating, repressing, and modulating posttranslational modifications. A comparison of the hTAF4-TAFH structure with the homologous ETO-TAFH domain reveals several critical residues important for hTAF4-TAFH target specificity and suggests that TAF4 has evolved in response to the increased transcriptional complexity of metazoans. TFIID | transcription | x-ray crystallography | TAFH domain.

Saccharomyces cerevisiae is an ideal host from which to obtain high levels of posttranslationally modified eukaryotic proteins for x-ray crystallography. However, extensive replacement of methionine by selenomethionine for anomalous dispersion phasing has proven intractable in yeast. We report a general method to incorporate selenomethionine into proteins expressed in yeast based on manipulation of the appropriate metabolic pathways, sam1- sam2mutants, in which the conversion of methionine to S-adenosylmethionine is blocked, exhibit reduced selenomethionine toxicity compared with wild-type yeast, increased production of protein during growth in selenomethionine, and efficient replacement of methionine by selenomethionine, based on quantitative mass spectrometry and x-ray crystallography. The structure of yeast tryptophanyl-tRNA synthetase was solved to 1.8 A by using multiwavelength anomalous dispersion phasing with protein that was expressed and purified from the sam1- sam2- strain grown in selenomethionine. Six of eight selenium residues were located in the structure. x-ray crystallography | methionine | Saccharomyces cerevisiae

The chlorination of bis(supermesityl)ditelluride RTeTeR (R = 2,4,6-t-[Bu.sub.3][C.sub.6][H.sub.6]) with 3 equiv of sulfuryl chloride S[O.sub.2][Cl.sub.2] has provided the unstable supermesityltellurium(IV) trichloride RTe[Cl.sub.3] as bright yellow crystals. X-ray crystallography is used to characterize the resulting compounds and depending on the aggregate state and the absence or presence of water, four distinctively different decomposition pathways are identified.

A dimolybdenum molecular pair, [Mo.sub.2][(DAniF).sub.3][([mu]-OH).sub.2][Mo.sub.2][(DAniF).sub.3], is synthesized and characterized by using X-ray crystallography. The results have provided an interesting case of single-crystal-to-single-crystal (SCSC) transformations and described the deprotonation process affected by dioxygen.

The solvatochromic behavior of hydroxyperylenebisimide was studied and the structure of naphthoperylenebisimide was determined by X-ray crystallography. The findings from the spectral studies indicate that the hydroxyperylenebisimide and naphthoperylenebisimide units of solvatochromic dyad were strongly coupled in the ground state and as a result the fluorescence of the naphthoperylenebisimide unit was almost quenched.

A computational study of species specificity for inhibitors of dihydrofolate reductase (DHFR) that focuses on incorporating protein flexibility into structure based drug-discovery is described. Models created for DHFR from human and Pneumocystis carinii incorporate a fair degree of protein flexibility and are highly selective for known DHFR inhibitors over drug-like non-inhibitors.

We determined the crystal structure of the Escherichia coli nucleoid-associated HU[alpha][beta] protein by x-ray diffraction and observed that the heterodimers form multimers with octameric units in three potential arrangements, which may serve specialized roles in different DNA transaction reactions. It is of special importance that one of the structures forms spiral filaments with left-handed rotations. A negatively superhelical DNA can be modeled to wrap around this lefthanded HU[alpha][beta] multimer. Whereas the wild-type HU generated negative DNA supercoiling in vitro, an engineered heterodimer with an altered amino acid residue critical for the formation of the left-handed spiral protein in the crystal was defective in the process, thus providing the structural explanation for the classical property of HU to restrain negative supercoils in DNA. HU-multimers | nucleoid | x-ray crystallography

In situ X-ray crystallographic observation is done to obtain the structural parameters of three persistent triplet diphenylcarbenes with significantly different stabilities. Significant differences are found between their structural information obtained in the crystalline state and that previously obtained spectroscopically in a glass matrix.

A study employs the X-ray crystallography technique to analyze the high-resolution crystal structure of a plant carboxylesterase, named Actinidia eriantha (AeCXEI), which is found to be closely related to the hormone-sensitive lipase subgroup structurally. The analysis reveals that the compound binds with paraoxon very easily forming a complex without inducing any change in the structure of the enzyme and hence helps in studying the role of carboxylesterases in plants.

A study analyzes the X-ray crystallographic and biochemical characterization of the action of an iimidazole-dioxolane compound on heme oxygenase (HO), which is a potential therapeutic target in cancer. The analysis reveals that the catalytic reactions of the HO complexes stop completely in the presence of the imidazole-dioxolane compound, as verdoheme has lower reactivity with oxygen as compared to the heme.

Zinc complexes of three new amide-appended ligands were prepared and isolated and the effect of replacement of the unsubstituted pyridyl groups in these complexes with various donors was studied. The results suggest that increased steric hindrance in the equatorial plane of the zinc cation slows or prevents the amide methanolysis reaction and the slower rate of amide methanolysis is due to a side reaction involving the formation of free ligand.

A new approach is described for the synthesis of macrocyclic tetralactones with different ring sizes having a wide variety of spacers. This method has also used Grubbs' catalyst for the synthesis of macrocyclic tetralactones having different ring sizes and spaces, which are important to the biologically significant materials and supramolecular chemistry.

The active sites of intramembrane proteases are positioned in the lipid bilayer to facilitate peptide bond hydrolysis in the membrane. Previous crystallographic analysis of Escherichia coli GlpG, an intramembrane protease of the rhomboid family, has revealed an internal and hydrophilic active site in an apparently closed conformation. Here we describe the crystal structure of GlpG in a more open conformation, where the capping loop L5 has been lifted, exposing the previously buried and catalytically essential Ser-201 to outside aqueous solution. A water molecule now moves into the putative oxyanion hole that is constituted of a main-chain amide (Ser-201) and two conserved side chains (His-150 and Asn-154). The loop movement also destabilizes a hydrophobic side chain (Phe-245) previously buried between transmembrane helices S2 and S5 and creates a side portal from the lipid to protease active site. These results provide insights into the conformational plasticity of GlpG to accommodate substrate binding and catalysis and into the chirality of the reaction intermediate. intramembrane proteolysis | membrane protein | rhomboid protease | x-ray crystallography

Ferrochelatase, the terminal enzyme in heme biosynthesis, catalyzes the insertion of ferrous iron into protoporphyrin IX to form protoheme IX. Human ferrochelatase is a homodimeric, inner mitochondrial membrane-associated enzyme that possesses an essential [2Fe-2S] cluster. In this work, we report the crystal structure of human ferrochelatase with the substrate protoporphyrin IX bound as well as a higher resolution structure of the R115L variant without bound substrate. The data presented reveal that the porphyrin substrate is bound deep within an enclosed pocket. When compared with the location of N-methylmesoporphyrin in the Bacillus subtilis ferrochelatase, the porphyrin is rotated by [approximately or equal to] 100[degrees] and is buried an additional 4.5 [Angstrom] deeper within the active site. The propionate groups of the substrate do not protrude into solvent and are bound in a manner similar to what has been observed in uroporphyrinogen decarboxylase. Furthermore, in the substrate-bound form, the jaws of the active site mouth are closed so that the porphyrin substrate is completely engulfed in the pocket. These data provide insights that will aid in the determination of the mechanism for ferrochelatase. heme biosynthesis | protoporphyrin IX | x-ray crystallography | metal insertion

Rhomboid peptidases are members of a family of regulated intramembrane peptidases that cleave the transmembrane segments of integral membrane proteins. Rhomboid peptidases have been shown to play a major role in developmental processes in Drosophila and in mitochondrial maintenance in yeast. Most recently, the function of rhomboid peptidases has been directly linked to apoptosis. We have solved the structure of the rhomboid peptidase from Haemophilus influenzae (hiGIpG) to 2.2-[Angstrom] resolution. The phasing for the crystals of hiGIpG was provided mainly by molecular replacement, by using the coordinates of the Escherichia coli rhomboid (ecGIpG). The structural results on these rhomboid peptidases have allowed us to speculate on the catalytic mechanism of substrate cleavage in a membranous environment. We have identified the relative disposition of the nucleophilic serine to the general base/acid function of the conserved histidine. Modeling a tetrapeptide substrate in the context of the rhomboid structure reveals an oxyanion hole comprising the side chain of a second conserved histidine and the main-chain NH of the nucleophilic serine residue. In both hiGIpG and ecGIpG structures, a water molecule occupies this oxyanion hole. intramembrane peptidase | membrane protein | rhomboid protease | x-ray crystallography

The solid-state structure and solution spectroscopic properties of the first example of a lanthanide tetrakis(dithiolene) complex along with the first examination of the redox chemistry between coordinated dithiolenes and a lanthanide ion are reported. This homoleptic lanthanide tetrakis are found to form a (dithiolene) complex forms a 2-D honeycomb network in the solid state due to its unusual dative N-Na bridging interactions from the nitrile termini of the mnt ligands.

The complete solid-state characterization and solution photophysics of several new complexes of lanthanide ions as possible sensitizers of Eu(III) and Tb(III) with thiophene-derivatized nitrobenzoic acids is reported. The compounds dissolve in water and methanol and methanol solutions were found to be luminescent which have metal ion-to-ligand ratio of 1:1.

HIV-1 protease is an effective target for designing drugs against AIDS, and structural information about the true transition state and the correct mechanism can provide important inputs. We present here the three-dimensional structure of a bi-product complex between HIV-1 protease and the two cleavage product peptides AETF and YVDGAA. The structure, refined against synchrotron data to 1.65 [Angstrom] resolution, shows the occurrence of the cleavage reaction in the crystal, with the product peptides still held in the enzyme active site. The separation between the scissile carbon and nitrogen atoms is 2.67 [Angstrom], which is shorter than a normal van der Waal separation, but it is much longer than a peptide bond length. The substrate is thus in a stage just past the G'Z intermediate described in Northrop's mechanism [Northrop DB (2001) Acc Chem Res 34:790-797]. Because the products are generated in situ, the structure, by extrapolation, can give insight into the mechanism of the cleavage reaction. Both oxygens of the generated carboxyl group form hydrogen bonds with atoms at the catalytic center: one to the OD2 atom of a catalytic aspartate and the other to the scissile nitrogen atom. The latter hydrogen bond may have mediated protonation of scissile nitrogen, triggering peptide bond cleavage. The inner oxygen atoms of the catalytic aspartates in the complex are 2.30 [Angstrom] apart, indicating a low-barrier hydrogen bond between them at this stage of the reaction, an observation not included in Northrop's proposal. This structure forms a template for designing mechanism-based inhibitors. AIDS | catalysis | reaction intermediate | x-ray crystallography

The syntheses, structural and spectroscopic characterization, fluoride abstraction reactions, and photochemical reactivity of cationic uranyl(VI) phosphine oxide complexes are described. The results illustrate an equal degree of lability associated with uranium(IV) oxidation as is found with uranyl(VI) reduction, presumably as both proceed through the same reactive uranyl(V) intermediate.