A 2.3 A resolution structure of chymosin complexed with a reduced bond inhibitor shows that the active site beta-hairpin flap is rearranged when compared with the native crystal structure

In the crystal structure of uncomplexed native chymosin, the beta- hairpin at the active site, known as 'the flap', adopts a different conformation from that of other aspartic proteinases. This conformation would prevent the mode of binding of substrates/inhibitors generally found in other aspartic proteinase complexes. We now report the X-ray analysis of chymosin complexed with a reduced bond inhibitor CP-113972 (2R,3S)-isopropyl 3-[(L-prolyl-p-iodo-L-phenylalanyl-S-methyl- cysteinyl)amino-4]-cyclohexy l-2-hydroxybutanoate at 2.3 A resolution in a novel crystal form of spacegroup R32. The structure has been refined by restrained least-squares methods to a final R-factor of 0.19 for a total of 11 988 independent reflections in the resolution range 10 to 2.3 A. The extended beta-strand conformation of the inhibitor allows hydrogen bonds within the active site, while its sidechains make both electrostatic and hydrophobic interactions with residues lining the specificity pockets S4-->S1. The flap closes over the active site cleft in a way that closely resembles that of other previously determined aspartic proteinase inhibitor complexes. We conclude that the usual position and conformation of the flap found in other aspartic proteinases is available to native chymosin. The conformation observed in the native crystal form may result from intermolecular interactions between symmetry-related molecules in the crystal lattice.      

Relative binding free energy calculations of inhibitors to two mutants (Glu46-- Ala/Gln) of ribonuclease T1 using molecular dynamics/free energy perturbation approaches

We present free energy perturbation calculations on the complexesof Glu46— Ala46 (E46A) and Glu46— Gln46 (E46Q) mutantsof ribonuclease T1 (RNaseT1) with inhibitors 2‘-guanosinemonophosphate (GMP) and 2’adenosine monophosphate (AMP)by a thermodynamic perturbation method implemented with moleculardynamics (MD). Using the available crystal structure of theRNaseT1–GMP complex, the structures of E46A-GMP and E46Q-GMPwere model built and equilibrated with MD simulations. The structuresof E46A-AMP and E46Q-AMP were obtained as a final structureof the GMP—AMP perturbation calculation respectively.The calculated difference in the free energy of binding (Gbind)was 0.31 kcal/mol for the E46A system and —1.04 kcal/molfor the E46Q system. The resultant free energies are much smallerthan the experimental and calculated value of 3 kcal/mol forthe native RNase T1, which suggests that both mutants have greaterrelative adenine affinities than native RNaseT1. EspeciallyE46Q is calculated to have a larger affinity for adenine thanguanine, as we suggested previously from the calculation onthe native RNaseT1. Thus, the molecular dynamics/free energyperturbation method may be helpful in protein engineering, directedtoward increasing or changing the substrate specificity of enzymes.     

Structural principles of the broad substrate specificity of Thermoactinomyces vulgaris carboxypeptidase T--role of amino acid residues at positions 260 and 262

An influence of residues at positions 260 and 262 on a broad substrate specificity of Thermoactinomyces vulgaris carboxypeptidase T (CPT) has been studied by means of site-directed mutagenesis. The structure of the S1'-site of CPT is similar to those of pancreatic carboxypeptidases A (CPA) and B (CPB); however, the enzyme is capable of cleaving off C-terminal hydrophobic (like CPA), C-terminal positively charged (like CPB), and negatively charged residues. The spatial alteration of the S1' site hydrophobic area in CPT by an insertion of one residue in the active site loop with Tyr255 by analogy with CPA and CPB did not change the enzyme specificity. The introduction of Ile262 (CPT D260G/T262I) led to a statistically significant reduction in activity towards charged substrates. The removal of a negative (CPT D260G) and placement of a positive charge (CPT D260G/T262K and CPT D260G/T262R) in the S1' site shifted the specificity of the variants towards substrates with C-terminal Glu. The selectivity profile was 64:1.7:1 for wild-type CPT, 815:115:1 for CPT D260G, 3270:1060:1 for CPT D260G/T262K and 1:2.4:0 for CPT D260G/T262R for substrates with C-terminal Leu, Glu and Arg, respectively. The obtained results confirm the important role of the amino acid residues at positions 260 and 262 in determination of the CPT substrate specificity.     

Computational studies on bergaptol O-methyltransferase from Ammi majus L.: The substrate specificity

In order to understand the mechanisms of substrate specificity and the interaction between bergaptol and bergaptol O-methyltransferase (BMT), a 3D model of BMT is generated based on the crystal structure of caffeic acid 3-O-methyltransferase (COMT EC 2.1.1.68, PDB code 1KYZ) by using the InsightII/Homology module. With the aid of the molecular mechanics and molecular dynamics methods, the final refined model is obtained and its reliability is further assessed by PROCHECK and ProSa2003. With this model, a flexible docking study is performed and the results indicate that BMT has narrow substrate specificity. Although the homology between both proteins is higher than 65% and all amino acids surrounding the binding site, except four residues, are similar in their sequences, the two proteins exhibit different substrate preferences. The differences in substrate specificity can be explained on the basis of the structures of the protein and the substrate. Our results indicate that His259 may be the catalytic base for the reaction, and Glu320, Glu287 bracket the catalytic His259. Especially, Glu320 forms a weak hydrogen bond with His259 and promotes transfer of an H ion.     

2,4,6-三硝基间二甲苯晶体的实验研究

合成了2,4,6-三硝基间二甲苯并培养出其晶体,用X射线单晶衍射,1HNMR,MS和傅里叶变换红外光谱进行了表征.该晶体为正交晶系,属Pbcn空间群,晶胞参数为a=5.749(2)×10-10 m,b=15.043(3)×10-10 m,c=11.415(2)×10-10 m, α=β=γ=90°, V=987.20(3)×10-30 m3, Z=4,Dc=1.623 g/cm3(理论密度),最终R因子为R1=0.0359, wR2=0.1006,所有数据的R因子为R1=0.0459, wR2=0.1045.依靠相邻分子中硝基氧之间的弱相相互作用,2,4,6-三硝基间二甲苯分子连接形成二维平面结构.对该晶体进行了DSC和TG-DTG热分析,根据热分析结果提出该物质经过一步失重过程完全分解.     

Macromolecular sorbent materials for urea capture

Three types of chitosan–glutaraldehyde (Chi–Glu) crosslinked copolymer materials were prepared at various Chi–Glu weight ratios (i.e., 1 : 0.0835, 1 : 0.334, and 1 : 0.585) and variable reaction times. The corresponding Chi–Glu copolymer materials were imbibed in CuSO₄ solution to yield impregnated materials in the form of copolymer/Cu(II) complexes. The copolymer materials were characterized using FTIR spectroscopy and thermogravimetry analysis. Urea sorption isotherms were obtained in aqueous solution at 295 K and pH 7 with pristine chitosan, Chi–Glu copolymers (i.e., 1 : 0.0835 and 1 : 0.585), and the corresponding Chi–Glu/Cu(II) complexes. The concentration of unbound urea was monitored indirectly using a colorimetric method with p‐dimethylaminobenzaldehyde. The equilibrium adsorption data were analyzed using the Sips isotherm model. The uptake of urea with pristine chitosan was 4.7% w/w, whereas Chi–Glu copolymers display increased sorption (Q_m = 10.6–17.1% w/w) with increasing glutaraldehyde content. Urea sorption is further enhanced (Q_m = 16.3–26.4% w/w) for copolymer Chi–Glu/Cu(II) complexes. The preparation of Chi–Glu copolymers at various conditions illustrates that the sorption capacity and molecular recognition of urea can be systematically tuned via crosslinking and the formation of copolymer/Cu(II) complexes, and these results are related to a previously reported study (Shimizu and Fujishige, J. Biomed. Mater. Res. 1983, 17, 597). © 2012 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013     

5-(4-叠氮呋咱基)-[1,2,3]三唑[4,5-c]并呋咱内盐的合成、晶体结构及性能

以3,4-二氨基呋咱为原料,经重氮化-叠氮化、氧化-环化等反应合成了一种新型无氢富氮含能材料5-(4-叠氮呋咱基)-[1,2,3]三唑[4,5-c]并呋咱内盐(AFTF);采用红外光谱、核磁共振、元素分析等方法表征了目标物的结构;获得了AFTF的单晶并进行了晶体结构解析;采用DSC方法研究了AFTF的热稳定性,初步探讨了氧化-环化反应机理;采用Gaussian 09程序CBS-QB3方法计算了AFTF的固相生成热,基于晶体密度和固相生成热,利用EXPLO5爆轰软件预估了AFTF的爆轰性能。结果表明,化合物AFTF晶体为正交晶系,空间群为P 2(1)2(1)2(1),晶胞参数为:a=8.1782(17),b=8.6446(18),c=11.521(2),V=814.5(3)3,Z=4,μ=0.151 mm^-1,F(000)=440;AFTF的熔点为101.02℃,热分解温度为186.39℃;AFTF晶体密度为1.795 g/cm 3(296 K),氮含量为63.6%,理论爆速为8.982 km/s,爆压为33.5 GPa,生成热为1178.9 kJ/mol,爆热为6450.8 kJ/kg,表明AFTF是一种爆轰性能优良的无氢富氮高能量密度化合物,有望应用于高能推进剂或气体发生剂领域;低熔点特性有望使其作为熔铸炸药载体使用。     

4,4‐Dimethyl‐ and Diastereomeric 4‐Hydroxy‐4‐methyl‐ (2S)‐Glutamate Analogues Display Distinct Pharmacological Profiles at Ionotropic Glutamate Receptors and Excitatory Amino Acid Transporters

Subtype‐selective ligands are of great interest to the scientific community, as they provide a tool for investigating the function of one receptor or transporter subtype when functioning in its native environment. Several 4‐substituted (S)‐glutamate (Glu) analogues were synthesized, and altogether this approach has provided important insight into the structure–activity relationships (SAR) for ionotropic and metabotropic glutamate receptors (iGluRs and mGluRs), as well as the excitatory amino acid transporters (EAATs). In this work, three 4,4‐disubstituted Glu analogues 1 – 3 , which are hybrid structures of important 4‐substituted Glu analogues 4 – 8 , were investigated at iGluRs and EAATs. Collectively, their pharmacological profiles add new and valuable information to the SAR for the iGluRs and EAAT1–3.     

Solubilizing Trityl-Type Tag To Synthesize Asx/Glx-Containing Peptides

A novel solubilizing tag system for Asp/Asn/Glu/Gln-containing peptides is described. In this method, an Asp/Glu[Dbz-Cys-NH₂]-containing peptide (Dbz: 3,4-diaminobenzoic acid) is first synthesized through fluorenylmethyloxycarbonyl (Fmoc) solid-phase peptide synthesis. The solubilizing moiety containing an oligo-Lys group is then attached to the peptide in hexafluoroisopropanol through a trityl anchor to afford a hydrophilic tagged peptide. To detach the solubilizing tag, the Dbz moiety of the tagged peptide is activated with NaNO₂, and the Asp/Asn/Glu/Gln-containing peptide is obtained through hydrolysis or ammonolysis. This synthetic approach proved to be compatible with native chemical ligation, and amyloid β-protein 1–42 was successfully synthesized by the solubilizing-tag-aided native chemical ligation–desulfurization method.     

Stabilization of the autoproteolysis of TNF-alpha converting enzyme (TACE) results in a novel crystal form suitable for structure-based drug design studies

The crystallization of TNF-alpha converting enzyme (TACE) has been useful in understanding the structure-activity relationships of new chemical entities. However, the propensity of TACE to undergo autoproteolysis has made enzyme handling difficult and impeded the identification of inhibitor soakable crystal forms. The autoproteolysis of TACE was found to be specific (Y352-V353) and occurred within a flexible loop that is in close proximity to the P-side of the active site. The rate of autoproteolysis was found to be proportional to the concentration of TACE, suggesting a bimolecular reaction mechanism. A limited specificity study of the S(1)' subsite was conducted using surrogate peptides and suggested substitutions that would stabilize the proteolysis of the loop at positions Y352-V353. Two mutant proteases (V353G and V353S) were generated and proved to be highly resistant to autoproteolysis. The kinetics of the more resistant mutant (V353G) and wild-type TACE were compared and demonstrated virtually identical IC(50) values for a panel of competitive inhibitors. However, the k(cat)/K(m) of the mutant for a larger substrate (P6 - P(6)') was approximately 5-fold lower than that for the wild-type enzyme. Comparison of the complexed wild-type and mutant structures indicated a subtle shift in a peripheral P-side loop (comprising the mutation site) that may be involved in substrate binding/turnover and might explain the mild kinetic difference. The characterization of this stabilized form of TACE has yielded an enzyme with similar native kinetic properties and identified a novel crystal form that is suitable for inhibitor soaking and structure determination.     

A new approach for correlating of H2S solubility in [emim][Lac], [bmim][ac] and [emim][pro] ionic liquids using two-parts combined models

Two bi-part combined models containing reaction equilibrium thermodynamic model and an equation of state (EoS) including cubic plus association (CPA) or modified Soave-Redlich-Kwong (SRK), were employed to correlate H₂S solubility in ionic liquids [emim][Lac], [bmim][Ac] and [emim][Pro]. RETM and CPA/mSRK are responsible for chemical and phase equilibria calculations, respectively. The RETM defines a complex formation reaction between an H₂S and two IL molecules into the liquid phase so that a complex in the form of H₂S(IL)₂ is probable to produce. On the other side, CPA/mSRK EoSs perform phase equilibria computations to find pressure and vapor phase concentrations. In the case of CPA, both of H₂S and the ionic liquids were considered as self-associating components which pursue 4C and 2B schemes, respectively. This model computes the pure IL parameters using experimental density data. In this work, critical properties were estimated through the modified Lydersen-Joback-Reid method, Eotvos and Guggenheim relations. Afterward, the binary systems were investigated by applying RETM. CPA presents average absolute deviations (AADs) equal to 2.41%, 13.42% and 3.52% for [emim][Lac], [bmim][Ac], and [emim][Pro], respectively. Moreover, the AADs obtained by mSRK are 3.75%, 5.07%, and 6.06%, respectively. As it is evident from the results, the combination of RETM with both CPA and mSRK EoSs will result in good correlation accuracy.     

Lamellar thickening behaviour of nylon-6,6 crystal by annealing

Nylon-6,6 — a typical polyamide — was annealed in the swollen state in glycerol to promote the partial melting of the polymer crystal. The recrystallization or lamellar thickening of nylon-6,6 crystal following partial melting was easily induced by this annealing, and the lamellar thickness of the crystal increased stepwise by $\text{1}\text{2}$ monomer unit length with increasing annealing temperature or annealing time. In addition, another distinct layer-thickening mechanism was observed which led to approximately doubling (and frequently quadrupling) the straight stem length of the lamellar crystal for all samples annealed under adequate conditions. New melting endotherms corresponding to these layer thicknesses (range of long spacings 140–180Å) were obtained by differential scanning calorimetry (d.s.c.) at temperatures ranging from 270° to 282°C. The mechanism of lamellar thickening is discussed with reference to the experimental results.     

二羟基苯二磺酸锰配合物的合成、晶体结构及热稳定性

在水溶剂中,采用回流法合成了两个二羟基苯二磺酸单核Mn(Ⅱ)含氮配体配合物Na[Mn(3,5-(SO3)2HCat)(phen)2H2O](1)和[Mn(3,5-(SO3)2H2Cat)(phen)2H2O]·H2O(2)(H2Cat=1,2-二羟基苯,phen=1,10-邻菲罗啉)。采用X射线单晶衍射、红外光谱、元素分析、紫外可见光谱和热重分析等方法对配合物进行了表征。X射线单晶衍射表征结果表明:两个配合物晶体均属于单斜晶系,晶族分别为P2(1)/c和P2(1)/n。两个配合物的配位方式相似,锰离子均与两个1,10-邻菲罗啉、一个水分子以及磺酸基配位。锰离子是六配位的。     

Crystal and molecular structure of an enantiomeric gossypol-acetic acid clathrate

Single crystals of gossypol with three molecules of acetic acid (gossypol triacetic acid) were grown from solutions of gossypol acetic acid and acetone. The crystals were unstable in air but could be stabilized for X-ray diffraction analysis by coating the crystal surfaces with a thin layer of mineral oil. The gossypol triacetic acid complex (C₃₀H₃₀O₈·3C₂H₄O₂) forms an orthorhombic crystal system with P2₁2₁2₁ (Z=4) symmetry. Unit cell dimensions were a=9.0208(7) Å, b=17.4884(10) Å, and c=24.358(2), Å yielding a volume of 3842.7(5) ų and a density of 1.2077(2) g/cm³. As with all previously reported crystals of gossypol, the gossypol molecules were of the aldehyde tautomer, and the two planar naphthalene rings were approximately perpendicular. Acetic acid molecules were found to lie in channels within the gossypol matrix. Individual crystals contained only one gossypol enantiomer, but both enantiomers crystallized from solution. Although the crystal habit could not be used to distinguish between the gossypol enantiomers, a fragment of the crystal could be derivatized and analyzed by high-performance liquid chromatography for this purpose. The ability to grow large, nonracemic crystals leads to a simple procedure for separating small quantities of the individual gossypol enantiomers.     

RG-CPA方程计算超临界CO2-醇体系汽液相平衡

综合考虑缔合作用和密度涨落的影响,将经重整化群修正的CPA方程（RG-CPA方程）扩展到二元体系汽液相平衡的计算。通过一个温度下的汽液相平衡数据回归得到二元交互作用参数,预测其他温度下的相平衡。采用这种方法计算了超临界CO₂与醇二元体系的汽液相平衡性质和临界曲线。结果表明,RG-CPA方程预测超临界CO₂与醇二元体系的气液相组成和密度具有较高的精度,液相组分平均绝对偏差为0.032,气相组分平均绝对偏差为0.019。与原始CPA方程相比,RG-CPA方程能更好地预测气液临界曲线。     

Electron Charge Density Distribution from X-ray Diffraction Study of the M-Nitrophenol Compound in the Monoclinic Form

At room temperature, the m-Nitrophenol (m-NPH) appears in two polymorphic structures: orthorhombic and monoclinic forms. In the present work, we shall focus on the monoclinic form of this compound which has a centrosymmetric structure with the space group P2₁/n. The molecular dipole moment has been estimated experimentally. High resolution single crystal diffraction experiment was performed at low temperature with MoKα radiation. The crystal structure was refined using the multipolar model of Hansen and Coppens (1978). The molecular electron charge density distribution is described accurately. The study reveals the nature of inter-molecular interactions including charge transfer and hydrogen bonds. In this crystal, hydrogen bonds of moderate strength occur between the hydroxyl group and the O atom in the nitro one.     

Effects of sintering temperature on microstructure and properties of low-grade bauxite-based ceramic proppant

Low density and high strength ceramic proppant was prepared by sintering high aluminium type low-grade bauxite and high iron type low-grade bauxite at the temperature range from 1300℃ to 1360℃. The phase composition and micromorphology of ceramic proppant were, respectively, characterized by X-ray diffraction (XRD) and scanning electron microscope (SEM). The bulk density, apparent density, and breakage ratio of ceramic proppant were tested using standard methods. The results show that mullite crystal and corundum crystal are fully developed, the spatial network structure of rod-shaped mullite is formed, and corundum crystal is evenly distributed in the sample when the sintering temperature is 1340℃. At this temperature, the bulk density of the sample is 1.42 g/cm³, the apparent density is 2.67 g/cm³, and the breakage ratio is 5.1% under the closure pressure of 52 MPa. Then, the growth mechanism of mullite crystal in ceramic proppant was explored. The results reveal that the mullite crystal in ceramic proppant conforms to the layer growth theory. The secondary crystal nucleus is formed in the growth process and shows a step growth mechanism. With the increase in sintering temperature, the preferred orientation growth of mullite crystal finally forms needle-like morphology.     

有效的[4+1]/[3+2+1]双环化反应构建茚并氧杂蒽杂环骨架及其晶体结构

茚并氧杂蒽类化合物C24H28O5是由邻苯二甲醛和5,5-二甲基-1,3-环己二酮在醋酸溶剂中,于室温下搅拌一步法反应制得,通过单晶X-射线衍射法测定其结构。该晶体属单斜晶系,空间群P2(1),晶胞参数a=1.006 92(11)nm,b=1.021 73(12)nm,c=2.430 6(2)nm,V=2.500 6(5)nm~3,Z=4,晶胞密度Dc=1.247 mg/m~3,吸收系数μ=0.087 mm~(-1),单胞中电子的数目F(000)=1 008,分子量Mr=469.56(单晶中包含一分子溶剂DMF)。晶体结构经全矩阵最小二乘法对各原子参数进行修正,最终得到的偏离因子为R=0.061 9,ωR=0.090 4。在晶体结构中新形成的茚环呈现信封式构象,同时形成的吡喃环显示为扭船式构象。     

含能配合物Ni（C5N5O5H-4）2（by）2的晶体结构和热分解性能

培养了含能配合物Ni(C5N5O5H42)(by2)(by=吡啶)晶体,用X射线单晶衍射法测定了其分子结构。其晶体属于单斜晶系,空间群为P21/C,a=9.0070(18)nm,b=10.002(2)nm,c=19.445(4)nm,β=93.69(3),V=1748.1(6)nm3,μ=0.633mm-1,Z=4,S=1.00,最终残差因子[I>2σ(I)]R1=0.0530,WR2=0.1162,对于全部数据R1=0.0965,WR2=0.1358。用DSC、TG-DTG对该配合物的热分解过程进行了研究。结果表明,该配合物的热分解过程仅由1个剧烈的放热峰组成,剩余残渣量约5.508%。用Kissinger法和Ozawa-Doyle法计算出配合物热分解过程中的表观活化能和指前因子分别为224.30kJ/mol和7.32×1019s-1。     

Gas Pressure Sintering of Silicon Nitride Powder Coated with Al2O3 and TiO2

Gas pressure sintering kinetics of silicon nitride powder coated with 10 wt% (9:1) Al₂O₃ and TiO₂ have been studied at 1850°C with a pressure schedule of 0.3 MPa in the first stage and 1 MPa in the second stage. The rates have been analyzed with a liquid-phase sintering model. Diffusion-controlled intermediate-stage kinetics have been observed. The role of second-step pressurization with nitrogen and argon has been determined by monitoring the kinetics. Pressurization at an earlier stage (∼90% relative density) reduces the densification rate but produces a denser material at the final stage. Although final density is greater, a porous surface layer forms on samples sintered with argon pressurization at the second stage. No such porous layer is formed in the case of pressurization with nitrogen. The mechanism of the intermediate-stage kinetics has been discussed with respect to the nature of the product analyzed by XRD after sintering.